Neisserial antigens

ABSTRACT

The invention provides proteins from  Neisseria meningitidis  (strains A &amp; B) and from  Neisseria gonorrhoeae , including amino acid sequences, the corresponding nucleotide sequences, expression data, and serological data. The proteins are useful antigens for vaccines, immunogenic compositions, and/or diagnostics.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a Divisional application of U.S. patent applicationSer. No. 12/653,954, filed, Dec. 18, 2009, which is a Divisional of U.S.patent application Ser. No. 10/864,684, filed Jun. 8, 2004, now U.S.Pat. No. 7,655,245, which is a continuation application of U.S. patentapplication Ser. No. 09/303,518, filed Apr. 30, 1999, now U.S. Pat. No.6,914,131, which is a continuation-in-part of International PatentApplication PCT/IB1998/001665, filed Oct. 9, 1998, from whichapplications priority is claimed pursuant to 35 U.S.C. §120.PCT/IB1998/001665 claims priority to Great Britain Patent ApplicationsNo. 9723516.2, filed Nov. 6, 1997; No. 9724190.5, filed Nov. 14, 1997;No. 9724386.9, filed Nov. 18, 1997; No. 9725158.1, filed Nov. 27, 1997;No. 9726147.3, filed Dec. 10, 1997; No. 9800759.4, filed Jan. 14, 1998;No. 9819016.8, filed Sep. 1, 1998. All of the above applications areincorporated herein by reference in their entirety.

SUBMISSION OF SEQUENCE LISTING ON ASCII TEXT FILE

The content of the following submission on ASCII text file isincorporated herein by reference in its entirety: a computer readableform (CRF) of the Sequence Listing (file name:223002099611SUBSEQLIST.txt, date recorded: Oct. 1, 2012, size: 2,099KB).

This invention relates to antigens from Neisseria bacteria.

BACKGROUND ART

Neisseria meningitidis and Neisseria gonorrhoeae are non-motile, gramnegative diplococci that are pathogenic in humans. N. meningitidiscolonises the pharynx and causes meningitis (and, occasionally,septicaemia in the absence of meningitis); N. gonorrhoeae colonises thegenital tract and causes gonorrhea. Although colonising different areasof the body and causing completely different diseases, the two pathogensare closely related, although one feature that clearly differentiatesmeningococcus from gonococcus is the presence of a polysaccharidecapsule that is present in all pathogenic meningococci.

N. gonorrhoeae caused approximately 800,000 cases per year during theperiod 1983-1990 in the United States alone (chapter by Meitzner &Cohen, “Vaccines Against Gonococcal Infection”, In: New GenerationVaccines, 2nd edition, ed. Levine, Woodrow, Kaper, & Cobon, MarcelDekker, New York, 1997, pp. 817-842). The disease causes significantmorbidity but limited mortality. Vaccination against N. gonorrhoeaewould be highly desirable, but repeated attempts have failed. The maincandidate antigens for this vaccine are surface-exposed proteins such aspili, porins, opacity-associated proteins (Opas) and othersurface-exposed proteins such as the Lip, Laz, IgA1 protease andtransferrin-binding proteins. The lipooligosaccharide (LOS) has alsobeen suggested as vaccine (Meitzner & Cohen, supra).

N. meningitidis causes both endemic and epidemic disease. In the UnitedStates the attack rate is 0.6-1 per 100,000 persons per year, and it canbe much greater during outbreaks (see Lieberman et al. (1996) Safety andImmunogenicity of a Serogroups A/C Neisseria meningitidisOligosaccharide-Protein Conjugate Vaccine in Young Children. JAMA275(19):1499-1503; Schuchat et al (1997) Bacterial Meningitis in theUnited States in 1995. N Engl J Med 337(14):970-976). In developingcountries, endemic disease rates are much higher and during epidemicsincidence rates can reach 500 cases per 100,000 persons per year.Mortality is extremely high, at 10-20% in the United States, and muchhigher in developing countries. Following the introduction of theconjugate vaccine against Haemophilus influenzae, N. meningitidis is themajor cause of bacterial meningitis at all ages in the United States(Schuchat et al (1997) supra).

Based on the organism's capsular polysaccharide, 12 serogroups of N.meningitidis have been identified. Group A is the pathogen most oftenimplicated in epidemic disease in sub-Saharan Africa. Serogroups B and Care responsible for the vast majority of cases in the United States andin most developed countries. Serogroups W135 and Y are responsible forthe rest of the cases in the United States and developed countries. Themeningococcal vaccine currently in use is a tetravalent polysaccharidevaccine composed of serogroups A, C, Y and W135. Although efficacious inadolescents and adults, it induces a poor immune response and shortduration of protection, and cannot be, used in infants [eg. Morbidityand Mortality weekly report, Vol. 46, No. RR-5 (1997)]. This is becausepolysaccharides are T-cell independent antigens that induce a weakimmune response that cannot be boosted by repeated immunization.Following the success of the vaccination against H. influenzae,conjugate vaccines against serogroups A and C have been developed andare at the final stage of clinical testing (Zollinger W D “New andImproved Vaccines Against Meningococcal Disease” in: New GenerationVaccines, supra, pp. 469-488; Lieberman et al (1996) supra; Costantinoet al (1992) Development and phase I clinical testing of a conjugatevaccine against meningococcus A and C. Vaccine 10:691-698).

Meningococcus B remains a problem, however. This serotype currently isresponsible for approximately 50% of total meningitis in the UnitedStates, Europe, and South America. The polysaccharide approach cannot beused because the menB capsular polysaccharide is a polymer ofα(2-8)-linked N-acetyl neuraminic acid that is also present in mammaliantissue. This results in tolerance to the antigen; indeed, if an immuneresponse were elicited, it would be anti-self, and thereforeundesirable. In order to avoid induction of autoimmunity and to induce aprotective immune response, the capsular polysaccharide has, forinstance, been chemically modified substituting the N-acetyl groups withN-propionyl groups, leaving the specific antigenicity unaltered (Romero& Outschoorn (1994) Current status of Meningococcal group B vaccinecandidates: capsular or non-capsular? Clin Microbiol Rev 7(4):559-575).

Alternative approaches to menB vaccines have used complex mixtures ofouter membrane proteins (OMPs), containing either the OMPs alone, orOMPs enriched in porins, or deleted of the class 4 OMPs that arebelieved to induce antibodies that block bactericidal activity. Thisapproach produces vaccines that are not well characterized. They areable to protect against the homologous strain, but are not effective atlarge where there are many antigenic variants of the outer membraneproteins. To overcome the antigenic variability, multivalent vaccinescontaining up to nine different porins have been constructed (eg.Poolman J T (1992) Development of a meningococcal vaccine. Infect.Agents Dis. 4:13-28). Additional proteins to be used in outer membranevaccines have been the opa and opc proteins, but none of theseapproaches have been able to overcome the antigenic variability (eg.Ala'Aldeen & Borriello (1996) The meningococcal transferrin-bindingproteins 1 and 2 are both surface exposed and generate bactericidalantibodies capable of killing homologous and heterologous strains.Vaccine 14(1):49-53).

A certain amount of sequence data is available for meningococcal andgonoccocal genes and proteins (eg. EP-A-0467714, WO96/29412), but thisis by no means complete. The provision of further sequences couldprovide an opportunity to identify secreted or surface-exposed proteinsthat are presumed targets for the immune system and which are notantigenically variable. For instance, some of the identified proteinscould be components of efficacious vaccines against meningococcus B,some could be components of vaccines against all meningococcalserotypes, and others could be components of vaccines against allpathogenic Neisseriae.

THE INVENTION

The invention provides proteins comprising the Neisserial amino acidsequences disclosed in the examples. These sequences relate to N.meningitidis or N. gonorrhoeae.

It also provides proteins comprising sequences homologous (ie. havingsequence identity) to the Neisserial amino acid sequences disclosed inthe examples. Depending on the particular sequence, the degree ofidentity is preferably greater than 50% (eg. 65%, 80%, 90%, or more).These homologous proteins include mutants and allelic variants of thesequences disclosed in the examples. Typically, 50% identity or morebetween two proteins is considered to be an indication of functionalequivalence. Identity between the proteins is preferably determined bythe Smith-Waterman homology search algorithm as implemented in theMPSRCH program (Oxford Molecular), using an affine gap search withparameters gap open penalty=12 and gap extension penalty=1.

The invention further provides proteins comprising fragments of theNeisserial amino acid sequences disclosed in the examples. The fragmentsshould comprise at least n consecutive amino acids from the sequencesand, depending on the particular sequence, n is 7 or more (eg. 8, 10,12, 14, 16, 18, 20 or more). Preferably the fragments comprise anepitope from the sequence.

The proteins of the invention can, of course, be prepared by variousmeans (eg. recombinant expression, purification from cell culture,chemical synthesis etc.) and in various forms (eg. native, fusionsetc.). They are preferably prepared in substantially pure or isolatedform (ie. substantially free from other Neisserial or host cellproteins)

According to a further aspect, the invention provides antibodies whichbind to these proteins. These may be polyclonal or monoclonal and may beproduced by any suitable means.

According to a further aspect, the invention provides nucleic acidcomprising the Neisserial nucleotide sequences disclosed in theexamples. In addition, the invention provides nucleic acid comprisingsequences homologous (ie. having sequence identity) to the Neisserialnucleotide sequences disclosed in the examples.

Furthermore, the invention provides nucleic acid which can hybridise tothe Neisserial nucleic acid disclosed in the examples, preferably under“high stringency” conditions (eg. 65° C. in a 0.1×SSC, 0.5% SDSsolution).

Nucleic acid comprising fragments of these sequences are also provided.These should comprise at least n consecutive nucleotides from theNeisserial sequences and, depending on the particular sequence, n is 10or more (eg 12, 14, 15, 18, 20, 25, 30, 35, 40 or more).

According to a further aspect, the invention provides nucleic acidencoding the proteins and protein fragments of the invention.

It should also be appreciated that the invention provides nucleic acidcomprising sequences complementary to those described above (eg. forantisense or probing purposes).

Nucleic acid according to the invention can, of course, be prepared inmany ways (eg. by chemical synthesis, from genomic or cDNA libraries,from the organism itself etc.) and can take various forms (eg. singlestranded, double stranded, vectors, probes etc.).

In addition, the term “nucleic acid” includes DNA and RNA, and alsotheir analogues, such as those containing modified backbones, and alsopeptide nucleic acids (PNA) etc.

According to a further aspect, the invention provides vectors comprisingnucleotide sequences of the invention (eg. expression vectors) and hostcells transformed with such vectors.

According to a further aspect, the invention provides compositionscomprising protein, antibody, and/or nucleic acid according to theinvention. These compositions may be suitable as vaccines, for instance,or as diagnostic reagents, or as immunogenic compositions.

The invention also provides nucleic acid, protein, or antibody accordingto the invention for use as medicaments (eg. as vaccines) or asdiagnostic reagents. It also provides the use of nucleic acid, protein,or antibody according to the invention in the manufacture of: (i) amedicament for treating or preventing infection due to Neisserialbacteria; (ii) a diagnostic reagent for detecting the presence ofNeisserial bacteria or of antibodies raised against Neisserial bacteria;and/or (iii) a reagent which can raise antibodies against Neisserialbacteria. Said Neisserial bacteria may be any species or strain (such asN. gonorrhoeae, or any strain of N. meningitidis, such as strain A,strain B or strain C).

The invention also provides a method of treating a patient, comprisingadministering to the patient a therapeutically effective amount ofnucleic acid, protein, and/or antibody according to the invention.

According to further aspects, the invention provides various processes.

A process for producing proteins of the invention is provided,comprising the step of culturing a host cell according to the inventionunder conditions which induce protein expression.

A process for producing protein or nucleic acid of the invention isprovided, wherein the protein or nucleic acid is synthesised in part orin whole using chemical means.

A process for detecting polynucleotides of the invention is provided,comprising the steps of: (a) contacting a nucleic probe according to theinvention with a biological sample under hybridizing conditions to formduplexes; and (b) detecting said duplexes.

A process for detecting proteins of the invention is provided,comprising the steps of: (a) contacting an antibody according to theinvention with a biological sample under conditions suitable for theformation of an antibody-antigen complexes; and (b) detecting saidcomplexes.

A summary of standard techniques and procedures which may be employed inorder to perform the invention (eg. to utilise the disclosed sequencesfor vaccination or diagnostic purposes) follows. This summary is not alimitation on the invention but, rather, gives examples that may beused, but are not required.

General

The practice of the present invention will employ, unless otherwiseindicated, conventional techniques of molecular biology, microbiology,recombinant DNA, and immunology, which are within the skill of the art.Such techniques are explained fully in the literature eg. SambrookMolecular Cloning; A Laboratory Manual, Second Edition (1989); DNACloning, Volumes I and ii (D. N Glover ed. 1985); OligonucleotideSynthesis (M. J. Gait ed, 1984); Nucleic Acid Hybridization (B. D. Hames& S. J. Higgins eds. 1984); Transcription and Translation (B. D. Hames &S. J. Higgins eds. 1984); Animal Cell Culture (R. I. Freshney ed. 1986);Immobilized Cells and Enzymes (IRL Press, 1986); B. Perbal, A PracticalGuide to Molecular Cloning (1984); the Methods in Enzymology series(Academic Press, Inc.), especially volumes 154 & 155; Gene TransferVectors for Mammalian Cells (J. H. Miller and M. P. Calm eds. 1987, ColdSpring Harbor Laboratory); Mayer and Walker, eds. (1987), ImmunochemicalMethods in Cell and Molecular Biology (Academic Press, London); Scopes,(1987) Protein Purification: Principles and Practice, Second Edition(Springer-Verlag, N.Y.), and Handbook of Experimental Immunology,Volumes I-IV (D. M. Weir and C. C. Blackwell eds 1986).

Standard abbreviations for nucleotides and amino acids are used in thisspecification.

All publications, patents, and patent applications cited herein areincorporated in full by reference. In particular, the contents of UKpatent applications 9723516.2, 9724190.5, 9724386.9, 9725158.1,9726147.3, 9800759.4, and 9819016.8 are incorporated herein.

DEFINITIONS

A composition containing X is “substantially free of” Y when at least85% by weight of the total X+Y in the composition is X. Preferably, Xcomprises at least about 90% by weight of the total of X+Y in thecomposition, more preferably at least about 95% or even 99% by weight.

The term “comprising” means “including” as well as “consisting” eg. acomposition “comprising” X may consist exclusively of X or may includesomething additional to X, such as X+Y.

The term “heterologous” refers to two biological components that are notfound together in nature. The components may be host cells, genes, orregulatory regions, such as promoters. Although the heterologouscomponents are not found together in nature, they can function together,as when a promoter heterologous to a gene is operably linked to thegene. Another example is where a Neisserial sequence is heterologous toa mouse host cell. A further examples would be two epitopes from thesame or different proteins which have been assembled in a single proteinin an arrangement not found in nature.

An “origin of replication” is a polynucleotide sequence that initiatesand regulates replication of polynucleotides, such as an expressionvector. The origin of replication behaves as an autonomous unit ofpolynucleotide replication within a cell, capable of replication underits own control. An origin of replication may be needed for a vector toreplicate in a particular host cell. With certain origins ofreplication, an expression vector can be reproduced at a high copynumber in the presence of the appropriate proteins within the cell.Examples of origins are the autonomously replicating sequences, whichare effective in yeast; and the viral T-antigen, effective in COS-7cells.

A “mutant” sequence is defined as DNA, RNA or amino acid sequencediffering from but having sequence identity with the native or disclosedsequence. Depending on the particular sequence, the degree of sequenceidentity between the native or disclosed sequence and the mutantsequence is preferably greater than 50% (eg. 60%, 70%, 80%, 90%, 95%,99% or more, calculated using the Smith-Waterman algorithm as describedabove). As used herein, an “allelic variant” of a nucleic acid molecule,or region, for which nucleic acid sequence is provided herein is anucleic acid molecule, or region, that occurs essentially at the samelocus in the genome of another or second isolate, and that, due tonatural variation caused by, for example, mutation or recombination, hasa similar but not identical nucleic acid sequence. A coding regionallelic variant typically encodes a protein having similar activity tothat of the protein encoded by the gene to which it is being compared.An allelic variant can also comprise an alteration in the 5′ or 3′untranslated regions of the gene, such as in regulatory control regions(eg. see U.S. Pat. No. 5,753,235).

Expression Systems

The Neisserial nucleotide sequences can be expressed in a variety ofdifferent expression systems; for example those used with mammaliancells, baculoviruses, plants, bacteria, and yeast.

i. Mammalian Systems

Mammalian expression systems are known in the art. A mammalian promoteris any DNA sequence capable of binding mammalian RNA polymerase andinitiating the downstream (3′) transcription of a coding sequence (eg.structural gene) into mRNA. A promoter will have a transcriptioninitiating region, which is usually placed proximal to the 5′ end of thecoding sequence, and a TATA box, usually located 25-30 base pairs (bp)upstream of the transcription initiation site. The TATA box is thoughtto direct RNA polymerase II to begin RNA synthesis at the correct site.A mammalian promoter will also contain an upstream promoter element,usually located within 100 to 200 bp upstream of the TATA box. Anupstream promoter element determines the rate at which transcription isinitiated and can act in either orientation [Sambrook et al. (1989)“Expression of Cloned Genes in Mammalian Cells.” In Molecular Cloning: ALaboratory Manual, 2nd ed.].

Mammalian viral genes are often highly expressed and have a broad hostrange; therefore sequences encoding mammalian viral genes provideparticularly useful promoter sequences. Examples include the SV40 earlypromoter, mouse mammary tumor virus LTR promoter, adenovirus major latepromoter (Ad MLP), and herpes simplex virus promoter. In addition,sequences derived from non-viral genes, such as the murinemetallotheionein gene, also provide useful promoter sequences.Expression may be either constitutive or regulated (inducible),depending on the promoter can be induced with glucocorticoid inhormone-responsive cells.

The presence of an enhancer element (enhancer), combined with thepromoter elements described above, will usually increase expressionlevels. An enhancer is a regulatory DNA sequence that can stimulatetranscription up to 1000-fold when linked to homologous or heterologouspromoters, with synthesis beginning at the normal RNA start site.Enhancers are also active when they are placed upstream or downstreamfrom the transcription initiation site, in either normal or flippedorientation, or at a distance of more than 1000 nucleotides from thepromoter [Maniatis et al. (1987) Science 236:1237; Alberts et al. (1989)Molecular Biology of the Cell, 2nd ed.]. Enhancer elements derived fromviruses may be particularly useful, because they usually have a broaderhost range. Examples include the SV40 early gene enhancer [Dijkema et al(1985) EMBO J. 4:761] and the enhancer/promoters derived from the longterminal repeat (LTR) of the Rous Sarcoma Virus [Gorman et al. (1982b)Proc. Natl. Acad. Sci. 79:6777] and from human cytomegalovirus [Boshartet al. (1985) Cell 41:521]. Additionally, some enhancers are regulatableand become active only in the presence of an inducer, such as a hormoneor metal ion [Sassone-Corsi and Borelli (1986) Trends Genet. 2:215;Maniatis et al. (1987) Science 236:1237].

A DNA molecule may be expressed intracellularly in mammalian cells. Apromoter sequence may be directly linked with the DNA molecule, in whichcase the first amino acid at the N-terminus of the recombinant proteinwill always be a methionine, which is encoded by the ATG start codon. Ifdesired, the N-terminus may be cleaved from the protein by in vitroincubation with cyanogen bromide.

Alternatively, foreign proteins can also be secreted from the cell intothe growth media by creating chimeric DNA molecules that encode a fusionprotein comprised of a leader sequence fragment that provides forsecretion of the foreign protein in mammalian cells. Preferably, thereare processing sites encoded between the leader fragment and the foreigngene that can be cleaved either in vivo or in vitro. The leader sequencefragment usually encodes a signal peptide comprised of hydrophobic aminoacids which direct the secretion of the protein from the cell. Theadenovirus triparite leader is an example of a leader sequence thatprovides for secretion of a foreign protein in mammalian cells.

Usually, transcription termination and polyadenylation sequencesrecognized by mammalian cells are regulatory regions located 3′ to thetranslation stop codon and thus, together with the promoter elements,flank the coding sequence. The 3′ terminus of the mature mRNA is formedby site-specific post-transcriptional cleavage and polyadenylation[Birnstiel et al. (1985) Cell 41:349; Proudfoot and Whitelaw (1988)“Termination and 3′ end processing of eukaryotic RNA. In Transcriptionand splicing (ed. B. D. Hames and D. M. Glover); Proudfoot (1989) TrendsBiochem. Sci. 14:105]. These sequences direct the transcription of anmRNA which can be translated into the polypeptide encoded by the DNA.Examples of transcription terminater/polyadenylation signals includethose derived from SV40 [Sambrook et al (1989) “Expression of clonedgenes in cultured mammalian cells.” In Molecular Cloning: A LaboratoryManual].

Usually, the above described components, comprising a promoter,polyadenylation signal, and transcription termination sequence are puttogether into expression constructs. Enhancers, introns with functionalsplice donor and acceptor sites, and leader sequences may also beincluded in an expression construct, if desired. Expression constructsare often maintained in a replicon, such as an extrachromosomal element(eg. plasmids) capable of stable maintenance in a host, such asmammalian cells or bacteria. Mammalian replication systems include thosederived from animal viruses, which require trans-acting factors toreplicate. For example, plasmids containing the replication systems ofpapovaviruses, such as SV40 [Gluzman (1981) Cell 23:175] orpolyomavirus, replicate to extremely high copy number in the presence ofthe appropriate viral T antigen. Additional examples of mammalianreplicons include those derived from bovine papillomavirus andEpstein-Barr virus. Additionally, the replicon may have two replicatonsystems, thus allowing it to be maintained, for example, in mammaliancells for expression and in a prokaryotic host for cloning andamplification. Examples of such mammalian-bacteria shuttle vectorsinclude pMT2 [Kaufman et al. (1989) Mol. Cell. Biol. 9:946] and pHEBO[Shimizu et al. (1986) Mol. Cell. Biol. 6:1074].

The transformation procedure used depends upon the host to betransformed. Methods for introduction of heterologous polynucleotidesinto mammalian cells are known in the art and include dextran-mediatedtransfection, calcium phosphate precipitation, polybrene mediatedtransfection, protoplast fusion, electroporation, encapsulation of thepolynucleotide(s) in liposomes, and direct microinjection of the DNAinto nuclei.

Mammalian cell lines available as hosts for expression are known in theart and include many immortalized cell lines available from the AmericanType Culture Collection (ATCC), including but not limited to, Chinesehamster ovary (CHO) cells, HeLa cells, baby hamster kidney (BHK) cells,monkey kidney cells (COS), human hepatocellular carcinoma cells (eg. HepG2), and a number of other cell lines.

ii. Baculovirus Systems

The polynucleotide encoding the protein can also be inserted into asuitable insect expression vector, and is operably linked to the controlelements within that vector. Vector construction employs techniqueswhich are known in the art. Generally, the components of the expressionsystem include a transfer vector, usually a bacterial plasmid, whichcontains both a fragment of the baculovirus genome, and a convenientrestriction site for insertion of the heterologous gene or genes to beexpressed; a wild type baculovirus with a sequence homologous to thebaculovirus-specific fragment in the transfer vector (this allows forthe homologous recombination of the heterologous gene in to thebaculovirus genome); and appropriate insect host cells and growth media.

After inserting the DNA sequence encoding the protein into the transfervector, the vector and the wild type viral genome are transfected intoan insect host cell where the vector and viral genome are allowed torecombine. The packaged recombinant virus is expressed and recombinantplaques are identified and purified. Materials and methods forbaculovirus/insect cell expression systems are commercially available inkit form from, inter alia, Invitrogen, San Diego Calif. (“MaxBac” kit).These techniques are generally known to those skilled in the art andfully described in Summers and Smith, Texas Agricultural ExperimentStation Bulletin No. 1555 (1987) (hereinafter “Summers and Smith”).

Prior to inserting the DNA sequence encoding the protein into thebaculovirus genome, the above described components, comprising apromoter, leader (if desired), coding sequence of interest, andtranscription termination sequence, are usually assembled into anintermediate transplacement construct (transfer vector). This constructmay contain a single gene and operably linked regulatory elements;multiple genes, each with its owned set of operably linked regulatoryelements; or multiple genes, regulated by the same set of regulatoryelements. Intermediate transplacement constructs are often maintained ina replicon, such as an extrachromosomal element (eg. plasmids) capableof stable maintenance in a host, such as a bacterium. The replicon willhave a replication system, thus allowing it to be maintained in asuitable host for cloning and amplification.

Currently, the most commonly used transfer vector for introducingforeign genes into AcNPV is pAc373. Many other vectors, known to thoseof skill in the art, have also been designed. These include, forexample, pVL985 (which alters the polyhedrin start codon from ATG toATT, and which introduces a BamHI cloning site 32 basepairs downstreamfrom the ATT; see Luckow and Summers, Virology (1989) 17:31.

The plasmid usually also contains the polyhedrin polyadenylation signal(Miller et al. (1988) Ann. Rev. Microbiol., 42:177) and a prokaryoticampicillin-resistance (amp) gene and origin of replication for selectionand propagation in E. coli.

Baculovirus transfer vectors usually contain a baculovirus promoter. Abaculovirus promoter is any DNA sequence capable of binding abaculovirus RNA polymerase and initiating the downstream (5′ to 3′)transcription of a coding sequence (eg. structural gene) into mRNA. Apromoter will have a transcription initiation region which is usuallyplaced proximal to the 5′ end of the coding sequence. This transcriptioninitiation region usually includes an RNA polymerase binding site and atranscription initiation site. A baculovirus transfer vector may alsohave a second domain called an enhancer, which, if present, is usuallydistal to the structural gene. Expression may be either regulated orconstitutive.

Structural genes, abundantly transcribed at late times in a viralinfection cycle, provide particularly useful promoter sequences.Examples include sequences derived from the gene encoding the viralpolyhedron protein, Friesen et al., (1986) “The Regulation ofBaculovirus Gene Expression,” in: The Molecular Biology of Baculoviruses(ed. Walter Doerfler); EPO Publ. Nos. 127 839 and 155 476; and the geneencoding the p10 protein, Vlak et al., (1988), J. Gen. Virol. 69:765.

DNA encoding suitable signal sequences can be derived from genes forsecreted insect or baculovirus proteins, such as the baculoviruspolyhedrin gene (Carbonell et al. (1988) Gene, 73:409). Alternatively,since the signals for mammalian cell posttranslational modifications(such as signal peptide cleavage, proteolytic cleavage, andphosphorylation) appear to be recognized by insect cells, and thesignals required for secretion and nuclear accumulation also appear tobe conserved between the invertebrate cells and vertebrate cells,leaders of non-insect origin, such as those derived from genes encodinghuman α-interferon, Maeda et al., (1985), Nature 315:592; humangastrin-releasing peptide, Lebacq-Verheyden et al., (1988), Molec. Cell.Biol. 8:3129; human IL-2, Smith et al., (1985) Proc. Nat'l Acad. Sci.USA, 82:8404; mouse IL-3, (Miyajima et al., (1987) Gene 58:273; andhuman glucocerebrosidase, Martin et al. (1988) DNA, 7:99, can also beused to provide for secretion in insects.

A recombinant polypeptide or polyprotein may be expressedintracellularly or, if it is expressed with the proper regulatorysequences, it can be secreted. Good intracellular expression of nonfusedforeign proteins usually requires heterologous genes that ideally have ashort leader sequence containing suitable translation initiation signalspreceding an ATG start signal. If desired, methionine at the N-terminusmay be cleaved from the mature protein by in vitro incubation withcyanogen bromide.

Alternatively, recombinant polyproteins or proteins which are notnaturally secreted can be secreted from the insect cell by creatingchimeric DNA molecules that encode a fusion protein comprised of aleader sequence fragment that provides for secretion of the foreignprotein in insects. The leader sequence fragment usually encodes asignal peptide comprised of hydrophobic amino acids which direct thetranslocation of the protein into the endoplasmic reticulum.

After insertion of the DNA sequence and/or the gene encoding theexpression product precursor of the protein, an insect cell host isco-transformed with the heterologous DNA of the transfer vector and thegenomic DNA of wild type baculovirus—usually by co-transfection. Thepromoter and transcription termination sequence of the construct willusually comprise a 2-5 kb section of the baculovirus genome. Methods forintroducing heterologous DNA into the desired site in the baculovirusvirus are known in the art. (See Summers and Smith supra; Ju et al.(1987); Smith et al., Mol. Cell. Biol. (1983) 3:2156; and Luckow andSummers (1989)). For example, the insertion can be into a gene such asthe polyhedrin gene, by homologous double crossover recombination;insertion can also be into a restriction enzyme site engineered into thedesired baculovirus gene. Miller et al., (1989), Bioessays 4:91. The DNAsequence, when cloned in place of the polyhedrin gene in the expressionvector, is flanked both 5′ and 3′ by polyhedrin-specific sequences andis positioned downstream of the polyhedrin promoter.

The newly formed baculovirus expression vector is subsequently packagedinto an infectious recombinant baculovirus. Homologous recombinationoccurs at low frequency (between about 1% and about 5%); thus, themajority of the virus produced after cotransfection is still wild-typevirus. Therefore, a method is necessary to identify recombinant viruses.An advantage of the expression system is a visual screen allowingrecombinant viruses to be distinguished. The polyhedrin protein, whichis produced by the native virus, is produced at very high levels in thenuclei of infected cells at late times after viral infection.Accumulated polyhedrin protein forms occlusion bodies that also containembedded particles. These occlusion bodies, up to 15 μm in size, arehighly refractile, giving them a bright shiny appearance that is readilyvisualized under the light microscope. Cells infected with recombinantviruses lack occlusion bodies. To distinguish recombinant virus fromwild-type virus, the transfection supernatant is plagued onto amonolayer of insect cells by techniques known to those skilled in theart. Namely, the plaques are screened under the light microscope for thepresence (indicative of wild-type virus) or absence (indicative ofrecombinant virus) of occlusion bodies. “Current Protocols inMicrobiology” Vol. 2 (Ausubel et al. eds) at 16.8 (Supp. 10, 1990);Summers and Smith, supra; Miller et al. (1989).

Recombinant baculovirus expression vectors have been developed forinfection into several insect cells. For example, recombinantbaculoviruses have been developed for, inter alis: Aedes aegypti,Autographa californica, Bombyx mori, Drosophila melanogaster, Spodopterafrugiperda, and Trichoplusia ni (WO 89/046699; Carbonell et al., (1985)J. Virol. 56:153; Wright (1986) Nature 321:718; Smith et al., (1983)Mol. Cell. Biol. 3:2156; and see generally, Fraser, et al. (1989) InVitro Cell. Dev. Biol. 25:225).

Cells and cell culture media are commercially available for both directand fusion expression of heterologous polypeptides in abaculovirus/expression system; cell culture technology is generallyknown to those skilled in the art. See, eg. Summers and Smith supra.

The modified insect cells may then be grown in an appropriate nutrientmedium, which allows for stable maintenance of the plasmid(s) present inthe modified insect host. Where the expression product gene is underinducible control, the host may be grown to high density, and expressioninduced. Alternatively, where expression is constitutive, the productwill be continuously expressed into the medium and the nutrient mediummust be continuously circulated, while removing the product of interestand augmenting depleted nutrients. The product may be purified by suchtechniques as chromatography, eg. HPLC, affinity chromatography, ionexchange chromatography, etc.; electrophoresis; density gradientcentrifugation; solvent extraction, or the like. As appropriate, theproduct may be further purified, as required, so as to removesubstantially any insect proteins which are also secreted in the mediumor result from lysis of insect cells, so as to provide a product whichis at least substantially free of host debris, eg. proteins, lipids andpolysaccharides.

In order to obtain protein expression, recombinant host cells derivedfrom the transformants are incubated under conditions which allowexpression of the recombinant protein encoding sequence. Theseconditions will vary, dependent upon the host cell selected. However,the conditions are readily ascertainable to those of ordinary skill inthe art, based upon what is known in the art.

iii. Plant Systems

There are many plant cell culture and whole plant genetic expressionsystems known in the art. Exemplary plant cellular genetic expressionsystems include those described in patents, such as: U.S. Pat. No.5,693,506; U.S. Pat. No. 5,659,122; and U.S. Pat. No. 5,608,143.Additional examples of genetic expression in plant cell culture has beendescribed by Zenk, Phytochemistry 30:3861-3863 (1991). Descriptions ofplant protein signal peptides may be found in addition to the referencesdescribed above in Vaulcombe et al., Mol. Gen. Genet. 209:33-40 (1987);Chandler et al., Plant Molecular Biology 3:407-418 (1984); Rogers, J.Biol. Chem. 260:3731-3738 (1985); Rothstein et al., Gene 55:353-356(1987); Whittier et al., Nucleic Acids Research 15:2515-2535 (1987);Wirsel et al., Molecular Microbiology 3:3-14 (1989); Yu et al., Gene122:247-253 (1992). A description of the regulation of plant geneexpression by the phytohormone, gibberellic acid and secreted enzymesinduced by gibberellic acid can be found in R. L. Jones and J.MacMillin, Gibberellins: in: Advanced Plant Physiology, Malcolm B.Wilkins, ed., 1984 Pitman Publishing Limited, London, pp. 21-52.References that describe other metabolically-regulated genes: Sheen,Plant Cell, 2:1027-1038 (1990); Maas et al., EMBO J. 9:3447-3452 (1990);Benkel and Hickey, Proc. Natl. Acad. Sci. 84:1337-1339 (1987)

Typically, using techniques known in the art, a desired polynucleotidesequence is inserted into an expression cassette comprising geneticregulatory elements designed for operation in plants. The expressioncassette is inserted into a desired expression vector with companionsequences upstream and downstream from the expression cassette suitablefor expression in a plant host. The companion sequences will be ofplasmid or viral origin and provide necessary characteristics to thevector to permit the vectors to move DNA from an original cloning host,such as bacteria, to the desired plant host. The basic bacterial/plantvector construct will preferably provide a broad host range prokaryotereplication origin; a prokaryote selectable marker; and, forAgrobacterium transformations, T DNA sequences forAgrobacterium-mediated transfer to plant chromosomes. Where theheterologous gene is not readily amenable to detection, the constructwill preferably also have a selectable marker gene suitable fordetermining if a plant cell has been transformed. A general review ofsuitable markers, for example for the members of the grass family, isfound in Wilmink and Dons, 1993, Plant Mol. Biol. Reptr, 11(2):165-185.

Sequences suitable for permitting integration of the heterologoussequence into the plant genome are also recommended. These might includetransposon sequences and the like for homologous recombination as wellas Ti sequences which permit random insertion of a heterologousexpression cassette into a plant genome. Suitable prokaryote selectablemarkers include resistance toward antibiotics such as ampicillin ortetracycline. Other DNA sequences encoding additional functions may alsobe present in the vector, as is known in the art.

The nucleic acid molecules of the subject invention may be included intoan expression cassette for expression of the protein(s) of interest.Usually, there will be only one expression cassette, although two ormore are feasible. The recombinant expression cassette will contain inaddition to the heterologous protein encoding sequence the followingelements, a promoter region, plant 5′ untranslated sequences, initiationcodon depending upon whether or not the structural gene comes equippedwith one, and a transcription and translation termination sequence.Unique restriction enzyme sites at the 5′ and 3′ ends of the cassetteallow for easy insertion into a pre-existing vector.

A heterologous coding sequence may be for any protein relating to thepresent invention. The sequence encoding the protein of interest willencode a signal peptide which allows processing and translocation of theprotein, as appropriate, and will usually lack any sequence which mightresult in the binding of the desired protein of the invention to amembrane. Since, for the most part, the transcriptional initiationregion will be for a gene which is expressed and translocated duringgermination, by employing the signal peptide which provides fortranslocation, one may also provide for translocation of the protein ofinterest. In this way, the protein(s) of interest will be translocatedfrom the cells in which they are expressed and may be efficientlyharvested. Typically secretion in seeds are across the aleurone orscutellar epithelium layer into the endosperm of the seed. While it isnot required that the protein be secreted from the cells in which theprotein is produced, this facilitates the isolation and purification ofthe recombinant protein.

Since the ultimate expression of the desired gene product will be in aeucaryotic cell it is desirable to determine whether any portion of thecloned gene contains sequences which will be processed out as introns bythe host's splicosome machinery. If so, site-directed mutagenesis of the“intron” region may be conducted to prevent losing a portion of thegenetic message as a false intron code, Reed and Maniatis, Cell41:95-105, 1985.

The vector can be microinjected directly into plant cells by use ofmicropipettes to mechanically transfer the recombinant DNA. Crossway,Mol. Gen. Genet, 202:179-185, 1985. The genetic material may also betransferred into the plant cell by using polyethylene glycol, Krens, etal., Nature, 296, 72-74, 1982. Another method of introduction of nucleicacid segments is high velocity ballistic penetration by small particleswith the nucleic acid either within the matrix of small beads orparticles, or on the surface, Klein, et al., Nature, 327, 70-73, 1987and Knudsen and Muller, 1991, Planta, 185:330-336 teaching particlebombardment of barley endosperm to create transgenic barley. Yet anothermethod of introduction would be fusion of protoplasts with otherentities, either minicells, cells, lysosomes or other fusiblelipid-surfaced bodies, Fraley, et al., Proc. Natl. Acad. Sci. USA, 79,1859-1863, 1982.

The vector may also be introduced into the plant cells byelectroporation. (Fromm et al., Proc. Natl. Acad. Sci. USA 82:5824,1985). In this technique, plant protoplasts are electroporated in thepresence of plasmids containing the gene construct. Electrical impulsesof high field strength reversibly permeabilize biomembranes allowing theintroduction of the plasmids. Electroporated plant protoplasts reformthe cell wall, divide, and form plant callus.

All plants from which protoplasts can be isolated and cultured to givewhole regenerated plants can be transformed by the present invention sothat whole plants are recovered which contain the transferred gene. Itis known that practically all plants can be regenerated from culturedcells or tissues, including but not limited to all major species ofsugarcane, sugar beet, cotton, fruit and other trees, legumes andvegetables. Some suitable plants include, for example, species from thegenera Fragaria, Lotus, Medicago, Onobrychis, Trifolium, Trigonella,Vigna, Citrus, Linum, Geranium, Manihot, Daucus, Arabidopsis, Brassica,Raphanus, Sinapis, Atropa, Capsicum, Datura, Hyoscyamus, Lycopersion,Nicotiana, Solanum, Petunia, Digitalis, Majorana, Cichorium, Helianthus,Lactuca, Bromus, Asparagus, Antirrhinum, Hererocallis, Nemesia,Pelargonium, Panicum, Pennisetum, Ranunculus, Senecio, Salpiglossis,Cucumis, Browaalia, Glycine, Lolium, Zea, Triticum, Sorghum, and Datura.

Means for regeneration vary from species to species of plants, butgenerally a suspension of transformed protoplasts containing copies ofthe heterologous gene is first provided. Callus tissue is formed andshoots may be induced from callus and subsequently rooted.Alternatively, embryo formation can be induced from the protoplastsuspension. These embryos germinate as natural embryos to form plants.The culture media will generally contain various amino acids andhormones, such as auxin and cytokinins. It is also advantageous to addglutamic acid and proline to the medium, especially for such species ascorn and alfalfa. Shoots and roots normally develop simultaneously.Efficient regeneration will depend on the medium, on the genotype, andon the history of the culture. If these three variables are controlled,then regeneration is fully reproducible and repeatable.

In some plant cell culture systems, the desired protein of the inventionmay be excreted or alternatively, the protein may be extracted from thewhole plant. Where the desired protein of the invention is secreted intothe medium, it may be collected. Alternatively, the embryos andembryoless-half seeds or other plant tissue may be mechanicallydisrupted to release any secreted protein between cells and tissues. Themixture may be suspended in a buffer solution to retrieve solubleproteins. Conventional protein isolation and purification methods willbe then used to purify the recombinant protein. Parameters of time,temperature pH, oxygen, and volumes will be adjusted through routinemethods to optimize expression and recovery of heterologous protein.

iv. Bacterial Systems

Bacterial expression techniques are known in the art. A bacterialpromoter is any DNA sequence capable of binding bacterial RNA polymeraseand initiating the downstream (3′) transcription of a coding sequence(eg. structural gene) into mRNA. A promoter will have a transcriptioninitiation region which is usually placed proximal to the 5′ end of thecoding sequence. This transcription initiation region usually includesan RNA polymerase binding site and a transcription initiation site. Abacterial promoter may also have a second domain called an operator,that may overlap an adjacent RNA polymerase binding site at which RNAsynthesis begins. The operator permits negative regulated (inducible)transcription, as a gene repressor protein may bind the operator andthereby inhibit transcription of a specific gene. Constitutiveexpression may occur in the absence of negative regulatory elements,such as the operator. In addition, positive regulation may be achievedby a gene activator protein binding sequence, which, if present isusually proximal (5′) to the RNA polymerase binding sequence. An exampleof a gene activator protein is the catabolite activator protein (CAP),which helps initiate transcription of the lac operon in Escherichia coli(E. coli) [Raibaud et al. (1984) Annu. Rev. Genet. 18:173]. Regulatedexpression may therefore be either positive or negative, thereby eitherenhancing or reducing transcription.

Sequences encoding metabolic pathway enzymes provide particularly usefulpromoter sequences. Examples include promoter sequences derived fromsugar metabolizing enzymes, such as galactose, lactose (lac) [Chang etal. (1977) Nature 198:1056], and maltose. Additional examples includepromoter sequences derived from biosynthetic enzymes such as tryptophan(trp) [Goeddel et al. (1980) Nuc. Acids Res. 8:4057; Yelverton et al.(1981) Nucl. Acids Res. 9:731; U.S. Pat. No. 4,738,921; EP-A-0036776 andEP-A-0121775]. The g-laotamase (bla) promoter system [Weissmann (1981)“The cloning of interferon and other mistakes.” In Interferon 3 (ed. I.Gresser)], bacteriophage lambda PL [Shimatake et al. (1981) Nature292:128] and T5 [U.S. Pat. No. 4,689,406] promoter systems also provideuseful promoter sequences.

In addition, synthetic promoters which do not occur in nature alsofunction as bacterial promoters. For example, transcription activationsequences of one bacterial or bacteriophage promoter may be joined withthe operon sequences of another bacterial or bacteriophage promoter,creating a synthetic hybrid promoter [U.S. Pat. No. 4,551,433]. Forexample, the tac promoter is a hybrid trp-lac promoter comprised of bothtrp promoter and lac operon sequences that is regulated by the lacrepressor [Amann et al. (1983) Gene 25:167; de Boer et al. (1983) Proc.Natl. Acad. Sci. 80:21].

Furthermore, a bacterial promoter can include naturally occurringpromoters of non-bacterial origin that have the ability to bindbacterial RNA polymerase and initiate transcription. A naturallyoccurring promoter of non-bacterial origin can also be coupled with acompatible RNA polymerase to produce high levels of expression of somegenes in prokaryotes. The bacteriophage T7 RNA polymerase/promotersystem is an example of a coupled promoter system [Studier et al. (1986)J. Mol. Biol. 189:113; Tabor et al., (1985) Proc Natl. Acad. Sci.82:1074]. In addition, a hybrid promoter can also be comprised of abacteriophage promoter and an E. coli operator region (EPO-A-0 267 851).

In addition to a functioning promoter sequence, an efficient ribosomebinding site is also useful for the expression of foreign genes inprokaryotes. In E. coli, the ribosome binding site is called theShine-Dalgarno (SD) sequence and includes an initiation codon (ATG) anda sequence 3-9 nucleotides in length located 3-11 nucleotides upstreamof the initiation codon [Shine et al. (1975) Nature 254:34]. The SDsequence is thought to promote binding of mRNA to the ribosome by thepairing of bases between the SD sequence and the 3′ and of E. coli 16SrRNA [Steitz et al: (1979) “Genetic signals and nucleotide sequences inmessenger RNA.” In Biological Regulation and Development: GeneExpression (ed. R. F. Goldberger)]. To express eukaryotic genes andprokaryotic genes with weak ribosome-binding site [Sambrook et al.(1989) “Expression of cloned genes in Escherichia coli.” In MolecularCloning: A Laboratory Manual].

A DNA molecule may be expressed intracellularly. A promoter sequence maybe directly linked with the DNA molecule, in which case the first aminoacid at the N-terminus will always be a methionine, which is encoded bythe ATG start codon. If desired, methionine at the N-terminus may becleaved from the protein by in vitro incubation with cyanogen bromide orby either in vivo on in vitro incubation with a bacterial methionineN-terminal peptidase (EPO-A-0 219 237).

Fusion proteins provide an alternative to direct expression. Usually, aDNA sequence encoding the N-terminal portion of an endogenous bacterialprotein, or other stable protein, is fused to the 5′ end of heterologouscoding sequences. Upon expression, this construct will provide a fusionof the two amino acid sequences. For example, the bacteriophage lambdacell gene can be linked at the 5′ terminus of a foreign gene andexpressed in bacteria. The resulting fusion protein preferably retains asite for a processing enzyme (factor Xa) to cleave the bacteriophageprotein from the foreign gene [Nagai et al. (1984) Nature 309:810].Fusion proteins can also be made with sequences from the lacZ [Jia etal. (1987) Gene 60:197], trpE [Allen et al. (1987) J. Biotechnol. 5:93;Makoff et al. (1989) J. Gen. Microbiol. 135:11], and Chey [EP-A-0 324647] genes. The DNA sequence at the junction of the two amino acidsequences may or may not encode a cleavable site. Another example is aubiquitin fusion protein. Such a fusion protein is made with theubiquitin region that preferably retains a site for a processing enzyme(eg. ubiquitin specific processing-protease) to cleave the ubiquitinfrom the foreign protein. Through this method, native foreign proteincan be isolated [Miller et al. (1989) Bio/Technology 7:698].

Alternatively, foreign proteins can also be secreted from the cell bycreating chimeric DNA molecules that encode a fusion protein comprisedof a signal peptide sequence fragment that provides for secretion of theforeign protein in bacteria [U.S. Pat. No. 4,336,336]. The signalsequence fragment usually encodes a signal peptide comprised ofhydrophobic amino acids which direct the secretion of the protein fromthe cell. The protein is either secreted into the growth media(gram-positive bacteria) or into the periplasmic space, located betweenthe inner and outer membrane of the cell (gram-negative bacteria).Preferably there are processing sites, which can be cleaved either invivo or in vitro encoded between the signal peptide fragment and theforeign gene.

DNA encoding suitable signal sequences can be derived from genes forsecreted bacterial proteins, such as the E. coli outer membrane proteingene (ompA) [Masui et al. (1983), in: Experimental Manipulation of GeneExpression; Ghrayeb et al. (1984) EMBO J. 3:2437] and the E. colialkaline phosphatase signal sequence (phoA) [Oka et al. (1985) Proc.Natl. Acad. Sci. 82:7212]. As an additional example, the signal sequenceof the alpha-amylase gene from various Bacillus strains can be used tosecrete heterologous proteins from B. subtilis [Palva et al. (1982)Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 244 042].

Usually, transcription termination sequences recognized by bacteria areregulatory regions located 3′ to the translation stop codon, and thustogether with the promoter flank the coding sequence. These sequencesdirect the transcription of an mRNA which can be translated into thepolypeptide encoded by the DNA. Transcription termination sequencesfrequently include DNA sequences of about 50 nucleotides capable offorming stem loop structures that aid in terminating transcription.Examples include transcription termination sequences derived from geneswith strong promoters, such as the irp gene in E. coli as well as otherbiosynthetic genes.

Usually, the above described components, comprising a promoter, signalsequence (if desired), coding sequence of interest, and transcriptiontermination sequence, are put together into expression constructs.Expression constructs are often maintained in a replicon, such as anextrachromosomal element (eg. plasmids) capable of stable maintenance ina host, such as bacteria. The replicon will have a replication system,thus allowing it to be maintained in a prokaryotic host either forexpression or for cloning and amplification. In addition, a replicon maybe either a high or low copy number plasmid. A high copy number plasmidwill generally have a copy number ranging from about 5 to about 200, andusually about 10 to about 150. A host containing a high copy numberplasmid will preferably contain at least about 10, and more preferablyat least about 20 plasmids. Either a high or low copy number vector maybe selected, depending upon the effect of the vector and the foreignprotein on the host.

Alternatively, the expression constructs can be integrated into thebacterial genome with an integrating vector. Integrating vectors usuallycontain at least one sequence homologous to the bacterial chromosomethat allows the vector to integrate. Integrations appear to result fromrecombinations between homologous DNA in the vector and the bacterialchromosome. For example, integrating vectors constructed with DNA fromvarious Bacillus strains integrate into the Bacillus chromosome (EP-A-0127 328). Integrating vectors may also be comprised of bacteriophage oftransposon sequences.

Usually, extrachromosomal and integrating expression constructs maycontain selectable markers to allow for the selection of bacterialstrains that have been transformed. Selectable markers can be expressedin the bacterial host and may include genes which render bacteriaresistant to drugs such as ampicillin, chloramphenicol, erythromycin,kanamycin (neomycin), and tetracycline [Davies et al. (1978) Annu. Rev.Microbiol. 32:469]. Selectable markers may also include biosyntheticgenes, such as those in the histidine, tryptophan, and leucinebiosynthetic pathways.

Alternatively, some of the above described components can be puttogether in transformation vectors. Transformation vectors are usuallycomprised of a selectable market that is either maintained in a repliconor developed into an integrating vector, as described above.

Expression and transformation vectors, either extra-chromosomalreplicons or integrating vectors, have been developed for transformationinto many bacteria. For example, expression vectors have been developedfor, inter alfa, the following bacteria: Bacillus subtilis [Palva et al.(1982) Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 036 259 and EP-A-0 063953; WO 84/04541], Escherichia coli [Shimatake et al. (1981) Nature292:128; Amann et al. (1985) Gene 40:183; Studier et al. (1986) J. Mol.Biol. 189:113; EP-A-0 036 776, EP-A-0 136 829 and EP-A-0 136 907],Streptococcus cremoris [Powell et al. (1988) Appl. Environ. Microbiol.54:655]; Streptococcus lividans [Powell et al. (1988) Appl. Environ.Microbiol. 54:655], Streptomyces lividans [U.S. Pat. No. 4,745,056].

Methods of introducing exogenous DNA into bacterial hosts are well-knownin the art, and usually include either the transformation of bacteriatreated with CaCl₂ or other agents, such as divalent cations and DMSO.DNA can also be introduced into bacterial cells by electroporation.Transformation procedures usually vary with the bacterial specie to betransformed. See eg. [Masson et al. (1989) FEMS Microbiol. Lett. 60:273;Palva et al. (1982) Proc. Natl. Acad. Sci. USA 79:5582; EP-A-0 036 259and EP-A-0 063 953; WO 84/04541, Bacillus], [Miller et al. (1988) Proc.Natl. Acad. Sci. 85:856; Wang et al. (1990) J. Bacteriol. 172:949,Campylobacter], [Cohen et al. (1973) Proc. Natl. Acad. Sci. 69:2110;Dower et al. (1988) Nucleic Acids Res. 16:6127; Kushner (1978) “Animproved method for transformation of Escherichia coli withColE1-derived plasmids. In Genetic Engineering: Proceedings of theInternational Symposium on Genetic Engineering (eds. H. W. Boyer and S.Nicosia); Mandel et al. (1970) J. Mol. Biol. 53:159; Taketo (1988)Biochim. Biophys. Acta 949:318; Escherichia], [Chassy et al. (1987) FEMSMicrobiol. Lett. 44:173 Lactobacillus]; [Fiedler et al. (1988) Anal.Biochem 170:38, Pseudomonas]; [Augustin et al. (1990) FEMS Microbiol.Lett. 66:203, Staphylococcus], [Barany et al. (1980) J. Bacteriol.144:698; Harlander (1987) “Transformation of Streptococcus lactis byelectroporation, in: Streptococcal Genetics (ed. J. Ferretti and R.Curtiss III); Perry et al. (1981) Infect. Immun. 32:1295; Powell et al.(1988) Appl. Environ. Microbiol. 54:655; Somkuti et al. (1987) Proc. 4thEvr. Cong. Biotechnology 1:412, Streptococcus].

v. Yeast Expression

Yeast expression systems are also known to one of ordinary skill in theart. A yeast promoter is any DNA sequence capable of binding yeast RNApolymerase and initiating the downstream (3′) transcription of a codingsequence (eg. structural gene) into mRNA. A promoter will have atranscription initiation region which is usually placed proximal to the5′ end of the coding sequence. This transcription initiation regionusually includes an RNA polymerase binding site (the “TATA Box”) and atranscription initiation site. A yeast promoter may also have a seconddomain called an upstream activator sequence (UAS), which, if present,is usually distal to the structural gene. The UAS permits regulated(inducible) expression. Constitutive expression occurs in the absence ofa UAS. Regulated expression may be either positive or negative, therebyeither enhancing or reducing transcription.

Yeast is a fermenting organism with an active metabolic pathway,therefore sequences encoding enzymes in the metabolic pathway provideparticularly useful promoter sequences. Examples include alcoholdehydrogenase (ADH) (EP-A-0 284 044), enolase, glucokinase,glucose-6-phosphate isomerase, glyceraldehyde-3-phosphate-dehydrogenase(GAP or GAPDH), hexokinase, phosphofructokinase, 3-phosphoglyceratemutase, and pyruvate kinase (PyK) (EPO-A-0 329 203). The yeast PHO5gene, encoding acid phosphatase, also provides useful promoter sequences[Myanohara et al. (1983) Proc. Natl. Acad. Sci. USA 80:1).

In addition, synthetic promoters which do not occur in nature alsofunction as yeast promoters. For example, UAS sequences of one yeastpromoter may be joined with the transcription activation region ofanother yeast promoter, creating a synthetic hybrid promoter. Examplesof such hybrid promoters include the ADH regulatory sequence linked tothe GAP transcription activation region (U.S. Pat. Nos. 4,876,197 and4,880,734). Other examples of hybrid promoters include promoters whichconsist of the regulatory sequences of either the ADH2, GAL4, GAL10, ORPHO5 genes, combined with the transcriptional activation region of aglycolytic enzyme gene such as GAP or PyK (EP-A-0 164 556). Furthermore,a yeast promoter can include naturally occurring promoters of non-yeastorigin that have the ability to bind yeast RNA polymerase and initiatetranscription. Examples of such promoters include, inter alia, [Cohen etal. (1980) Proc. Natl. Acad. Sci. USA 77:1078; Henikoff et al. (1981)Nature 283:835; Hollenberg et al. (1981) Curr. Topics Microbiol.Immunol. 96:119; Hollenberg et al. (1979) “The Expression of BacterialAntibiotic Resistance Genes in the Yeast Saccharomyces cerevisiae,” in:Plasmids of Medical, Environmental and Commercial Importance (eds. K. N.Timmis and A. Puhler); Mercerau-Puigalon et al. (1980) Gene 11:163;Panthier et al. (1980) Curr. Genet. 2:109;].

A DNA molecule may be expressed intracellularly in yeast. A promotersequence may be directly linked with the DNA molecule, in which case thefirst amino acid at the N-terminus of the recombinant protein willalways be a methionine, which is encoded by the ATG start codon. Ifdesired, methionine at the N-terminus may be cleaved from the protein byin vitro incubation with cyanogen bromide.

Fusion proteins provide an alternative for yeast expression systems, aswell as in mammalian, baculovirus, and bacterial expression systems.Usually, a DNA sequence encoding the N-terminal portion of an endogenousyeast protein, or other stable protein, is fused to the 5′ end ofheterologous coding sequences. Upon expression, this construct willprovide a fusion of the two amino acid sequences. For example, the yeastor human superoxide dismutase (SOD) gene, can be linked at the 5′terminus of a foreign gene and expressed in yeast. The DNA sequence atthe junction of the two amino acid sequences may or may not encode acleavable site. See eg. EP-A-0 196 056. Another example is a ubiquitinfusion protein. Such a fusion protein is made with the ubiquitin regionthat preferably retains a site for a processing enzyme (eg,ubiquitin-specific processing protease) to cleave the ubiquitin from theforeign protein. Through this method, therefore, native foreign proteincan be isolated (eg. WO88/024066).

Alternatively, foreign proteins can also be secreted from the cell intothe growth media by creating chimeric DNA molecules that encode a fusionprotein comprised of a leader sequence fragment that provide forsecretion in yeast of the foreign protein. Preferably, there areprocessing sites encoded between the leader fragment and the foreigngene that can be cleaved either in vivo or in vitro. The leader sequencefragment usually encodes a signal peptide comprised of hydrophobic aminoacids which direct the secretion of the protein from the cell.

DNA encoding suitable signal sequences can be derived from genes forsecreted yeast proteins, such as the yeast invertase gene (EP-A-0 012873; JPO. 62,096,086) and the A-factor gene (U.S. Pat. No. 4,588,684).Alternatively, leaders of non-yeast origin, such as an interferonleader, exist that also provide for secretion in yeast (EP-A-0 060 057).

A preferred class of secretion leaders are those that employ a fragmentof the yeast alpha-factor gene, which contains both a “pre” signalsequence, and a “pro” region. The types of alpha-factor fragments thatcan be employed include the full-length pre-pro alpha factor leader(about 83 amino acid residues) as well as truncated alpha-factor leaders(usually about 25 to about 50 amino acid residues) (U.S. Pat. Nos.4,546,083 and 4,870,008; EP-A-0 324 274). Additional leaders employingan alpha-factor leader fragment that provides for secretion includehybrid alpha-factor leaders made with a presequence of a first yeast,but a pro-region from a second yeast alphafactor. (eg. see WO 89/02463.)

Usually, transcription termination sequences recognized by yeast areregulatory regions located 3′ to the translation stop codon, and thustogether with the promoter flank the coding sequence. These sequencesdirect the transcription of an mRNA which can be translated into thepolypeptide encoded by the DNA. Examples of transcription terminatorsequence and other yeast-recognized termination sequences, such as thosecoding for glycolytic enzymes.

Usually, the above described components, comprising a promoter, leader(if desired), coding sequence of interest, and transcription terminationsequence, are put together into expression constructs. Expressionconstructs are often maintained in a replicon, such as anextrachromosomal element (eg. plasmids) capable of stable maintenance ina host, such as yeast or bacteria. The repl icon may have tworeplication systems, thus allowing it to be maintained, for example, inyeast for expression and in a prokaryotic host for cloning andamplification. Examples of such yeast-bacteria shuttle vectors includeYEp24 [Botstein et al. (1979) Gene 8:17-24], pCl/1 [Brake et al. (1984)Proc. Natl. Acad. Sci. USA 81:4642-4646], and YRp17 [Stinchcomb et al.(1982) J. Mol. Biol. 158:157]. In addition, a replicon may be either ahigh or low copy number plasmid. A high copy number plasmid willgenerally have a copy number ranging from about 5 to about 200, andusually about 10 to about 150. A host containing a high copy numberplasmid will preferably have at least about 10, and more preferably atleast about 20. Enter a high or low copy number vector may be selected,depending upon the effect of the vector and the foreign protein on thehost. See eg. Brake et al., supra.

Alternatively, the expression constructs can be integrated into theyeast genome with an integrating vector. Integrating vectors usuallycontain at least one sequence homologous to a yeast chromosome thatallows the vector to integrate, and preferably contain two homologoussequences flanking the expression construct. Integrations appear toresult from recombinations between homologous DNA in the vector and theyeast chromosome [Orr-Weaver et al. (1983) Methods in Enzymol.101:228-245]. An integrating vector may be directed to a specific locusin yeast by selecting the appropriate homologous sequence for inclusionin the vector. See Orr-Weaver et al., supra. One or more expressionconstruct may integrate, possibly affecting levels of recombinantprotein produced [Rine et al. (1983) Proc. Natl. Acad. Sci. USA80:6750]. The chromosomal sequences included in the vector can occureither as a single segment in the vector, which results in theintegration of the entire vector, or two segments homologous to adjacentsegments in the chromosome and flanking the expression construct in thevector, which can result in the stable integration of only theexpression construct.

Usually, extrachromosomal and integrating expression constructs maycontain selectable markers to allow for the selection of yeast strainsthat have been transformed. Selectable markers may include biosyntheticgenes that can be expressed in the yeast host, such as ADE2, HIS4, LEU2,TRP1, and ALG7, and the G418 resistance gene, which confer resistance inyeast cells to tunicamycin and G418, respectively. In addition, asuitable selectable marker may also provide yeast with the ability togrow in the presence of toxic compounds, such as metal. For example, thepresence of CUP1 allows yeast to grow in the presence of copper ions[Butt et al. (1987) Microbiol, Rev. 51:351].

Alternatively, some of the above described components can be puttogether into transformation vectors. Transformation vectors are usuallycomprised of a selectable marker that is either maintained in a repliconor developed into an integrating vector, as described above.

Expression and transformation vectors, either extrachromosomal repliconsor integrating vectors, have been developed for transformation into manyyeasts. For example, expression vectors have been developed for, interalia, the following yeasts: Candida albicans [Kurtz, et al. (1986) Mol.Cell. Biol. 6:142], Candida maltosa [Kunze, et al. (1985) J. BasicMicrobiol. 25:141]. Hansenula polymorpha [Gleeson, et al. (1986) J. Gen.Microbiol. 132:3459; Roggenkamp et al. (1986) Mol. Gen. Genet. 202:302],Kluyveromyces fragilis [Das, et al. (1984) J. Bacteriol. 158:1165],Kluyveromyces lactis [De Louvencourt et al. (1983) J. Bacteriol.154:737; Van den Berg et al. (1990) Bio/Technology 8:135], Pichiaguillerimondii [Kunze et al. (1985) J. Basic Microbiol. 25:141], Pichiapastoris [Cregg, et al. (1985) Mol. Cell. Biol. 5:3376; U.S. Pat. Nos.4,837,148 and 4,929,555], Saccharomyces cerevisiae [Hinnen et al. (1978)Proc. Natl. Acad. Sci. USA 75:1929; Ito et al. (1983) J. Bacteriol.153:163], Schizosaccharomyces pombe [Beach and Nurse (1981) Nature300:706], and Yarrowia lipolytica [Davidow, et al. (1985) Curr. Genet.10:380471 Gaillardin, et al. (1985) Curr. Genet. 10:49].

Methods of introducing exogenous DNA into yeast hosts are well-known inthe art, and usually include either the transformation of spheroplastsor of intact yeast cells treated with alkali cations. Transformationprocedures usually vary with the yeast species to be transformed. Seeeg. [Kurtz et al. (1986) Mol. Cell. Biol. 6:142; Kunze et al. (1985) J.Basic Microbiol. 25:141; Candida]; [Gleeson et al. (1986) J. Gen.Microbiol. 132:3459; Roggenkamp et al. (1986) Mol. Gen. Genet. 202:302;Hansenula]; [Das et al. (1984) J. Bacteriol. 158:1165; De Louvencourt etal. (1983) J. Bacteriol. 154:1165; Van den Berg et al. (1990)Bio/Technology 8:135; Kluyveromyces]; [Cregg et al. (1985) Mol. Cell.Biol. 5:3376; Kunze et al. (1985) J. Basic Microbiol. 25:141; U.S. Pat.Nos. 4,837,148 and 4,929,555; Pichia]; [Hinnen et al. (1978) Proc. Natl.Acad. Sci. USA 75; 1929; Ito et al. (1983) J. Bacteriol. 153:163Saccharomyces]; [Beach and Nurse (1981) Nature 300:706;Schizosaccharomyces]; [Davidow et al. (1985) Curr. Genet. 10:39;Gaillardin et al. (1985) Curr. Genet. 10:49; Yarrowia].

Antibodies

As used herein, the term “antibody” refers to a polypeptide or group ofpolypeptides composed of at least one antibody combining site. An“antibody combining site” is the three-dimensional binding space with aninternal surface shape and charge distribution complementary to thefeatures of an epitope of an antigen, which allows a binding of theantibody with the antigen. “Antibody” includes, for example, vertebrateantibodies, hybrid antibodies, chimeric antibodies, humanisedantibodies, altered antibodies, univalent antibodies, Fab proteins, andsingle domain antibodies.

Antibodies against the proteins of the invention are useful for affinitychromatography, immunoassays, and distinguishing/identifying Neisserialproteins.

Antibodies to the proteins of the invention, both polyclonal andmonoclonal, may be prepared by conventional methods. In general, theprotein is first used to immunize a suitable animal, preferably a mouse,rat, rabbit or goat. Rabbits and goats are preferred for the preparationof polyclonal sera due to the volume of serum obtainable, and theavailability of labeled anti-rabbit and anti-goat antibodies.Immunization is generally performed by mixing or emulsifying the proteinin saline, preferably in an adjuvant such as Freund's complete adjuvant,and injecting the mixture or emulsion parenterally (generallysubcutaneously or intramuscularly): A dose of 50-200 μg/injection istypically sufficient. Immunization is generally boosted 2-6 weeks laterwith one or more injections of the protein in saline, preferably usingFreund's incomplete adjuvant. One may alternatively generate antibodiesby in vitro immunization using methods known in the art, which for thepurposes of this invention is considered equivalent to in vivoimmunization. Polyclonal antisera is obtained by bleeding the immunizedanimal into a glass or plastic container, incubating the blood at 25° C.for one hour, followed by incubating at 4° C. for 2-18 hours. The serumis recovered by centrifugation (eg. 1,000 g for 10 minutes). About 20-50ml per bleed may be obtained from rabbits.

Monoclonal antibodies are prepared using the standard method of Kohler &Milstein [Nature (1975) 256:495-96], or a modification thereof.Typically, a mouse or rat is immunized as described above. However,rather than bleeding the animal to extract serum, the spleen (andoptionally several large lymph nodes) is removed and dissociated intosingle cells. If desired, the spleen cells may be screened (afterremoval of nonspecifically adherent cells) by applying a cell suspensionto a plate or well coated with the protein antigen. B-cells expressingmembrane-bound immunoglobulin specific for the antigen bind to theplate, and are not rinsed away with the rest of the suspension.Resulting B-cells, or all dissociated spleen cells, are then induced tofuse with myeloma cells to form hybridomas, and are cultured in aselective medium (eg. hypoxanthine, aminopterin, thymidine medium,“HAT”). The resulting hybridomas are plated by limiting dilution, andare assayed for the production of antibodies which bind specifically tothe immunizing antigen (and which do not bind to unrelated antigens).The selected MAb-secreting hybridomas are then cultured either in vitro(eg. in tissue culture bottles or hollow fiber reactors), or in vivo (asascites in mice).

If desired, the antibodies (whether polyclonal or monoclonal) may belabeled using conventional techniques. Suitable labels includefluorophores, chromophores, radioactive atoms (particularly ³²P and¹²⁵I), electron-dense reagents, enzymes, and ligands having specificbinding partners. Enzymes are typically detected by their activity. Forexample, horseradish peroxidase is usually detected by its ability toconvert 3,3′,5,5′-tetramethylbenzidine (TMB) to a blue pigment,quantifiable with a spectrophotometer. “Specific binding partner” refersto a protein capable of binding a ligand molecule with high specificity,as for example in the case of an antigen and a monoclonal antibodyspecific therefor. Other specific binding partners include biotin andavidin or streptavidin, IgG and protein A, and the numerousreceptor-ligand couples known in the art. It should be understood thatthe above description is not meant to categorize the various labels intodistinct classes, as the same label may serve in several differentmodes. For example, ¹²⁵I may serve as a radioactive label or as anelectron-dense reagent. HRP may serve as enzyme or as antigen for a MAb.Further, one may combine various labels for desired effect. For example,MAbs and avidin also require labels in the practice of this invention:thus, one might label a MAb with biotin, and detect its presence withavidin labeled with ¹²⁵I, or with an anti-biotin MAb labeled with HRP.Other permutations and possibilities will be readily apparent to thoseof ordinary skill in the art, and are considered as equivalents withinthe scope of the instant invention.

Pharmaceutical Compositions

Pharmaceutical compositions can comprise either polypeptides,antibodies, or nucleic acid of the invention. The pharmaceuticalcompositions will comprise a therapeutically effective amount of eitherpolypeptides, antibodies, or polynucleotides of the claimed invention.

The term “therapeutically effective amount” as used herein refers to anamount of a therapeutic agent to treat, ameliorate, or prevent a desireddisease or condition, or to exhibit a detectable therapeutic orpreventative effect. The effect can be detected by, for example,chemical markers or antigen levels. Therapeutic effects also includereduction in physical symptoms, such as decreased body temperature. Theprecise effective amount for a subject will depend upon the subject'ssize and health, the nature and extent of the condition, and thetherapeutics or combination of therapeutics selected for administration.Thus, it is not useful to specify an exact effective amount in advance.However, the effective amount for a given situation can be determined byroutine experimentation and is within the judgement of the clinician.

For purposes of the present invention, an effective dose will be fromabout 0.01 mg/kg to 50 mg/kg or 0.05 mg/kg to about 10 mg/kg of the DNAconstructs in the individual to which it is administered.

A pharmaceutical composition can also contain a pharmaceuticallyacceptable carrier. The term “pharmaceutically acceptable carrier”refers to a carrier for administration of a therapeutic agent, such asantibodies or a polypeptide, genes, and other therapeutic agents. Theterm refers to any pharmaceutical carrier that does not itself inducethe production of antibodies harmful to the individual receiving thecomposition, and which may be administered without undue toxicity.Suitable carriers may be large, slowly metabolized macromolecules suchas proteins, polysaccharides, polylactic acids, polyglycolic acids,polymeric amino acids, amino acid copolymers, and inactive virusparticles. Such carriers are well known to those of ordinary skill inthe art.

Pharmaceutically acceptable salts can be used therein, for example,mineral acid salts such as hydrochlorides, hydrobromides, phosphates,sulfates, and the like; and the salts of organic acids such as acetates,propionates, malonates, benzoates, and the like. A thorough discussionof pharmaceutically acceptable excipients is available in Remington'sPharmaceutical Sciences (Mack Pub. Co., N.J. 1991).

Pharmaceutically acceptable carriers in therapeutic compositions maycontain liquids such as water, saline, glycerol and ethanol.Additionally, auxiliary substances, such as wetting or emulsifyingagents, pH buffering substances, and the like, may be present in suchvehicles. Typically, the therapeutic compositions are prepared asinjectables, either as liquid solutions or suspensions; solid formssuitable for solution in, or suspension in, liquid vehicles prior toinjection may also be prepared. Liposomes are included within thedefinition of a pharmaceutically acceptable carrier.

Delivery Methods

Once formulated, the compositions of the invention can be administereddirectly to the subject. The subjects to be treated can be animals; inparticular, human subjects can be treated.

Direct delivery of the compositions will generally be accomplished byinjection, either subcutaneously, intraperitoneally, intravenously orintramuscularly or delivered to the interstitial space of a tissue. Thecompositions can also be administered into a lesion. Other modes ofadministration include oral and pulmonary administration, suppositories,and transdermal or transcutaneous applications (eg. see WO98/20734),needles, and gene guns or hyposprays. Dosage treatment may be a singledose schedule or a multiple dose schedule.

Vaccines

Vaccines according to the invention may either be prophylactic (ie. toprevent infection) or therapeutic (ie. to treat disease afterinfection).

Such vaccines comprise immunising antigen(s), immunogen(s),polypeptide(s), protein(s) or nucleic acid, usually in combination with“pharmaceutically acceptable carriers,” which include any carrier thatdoes not itself induce the production of antibodies harmful to theindividual receiving the composition. Suitable carriers are typicallylarge, slowly metabolized macromolecules such as proteins,polysaccharides, polylactic acids, polyglycolic acids, polymeric aminoacids, amino acid copolymers, lipid aggregates (such as oil droplets orliposomes), and inactive virus particles. Such carriers are well knownto those of ordinary skill in the art. Additionally, these carriers mayfunction as immunostimulating agents (“adjuvants”). Furthermore, theantigen or immunogen may be conjugated to a bacterial toxoid, such as atoxoid from diphtheria, tetanus, cholera, H. pylori, etc. pathogens.

Preferred adjuvants to enhance effectiveness of the composition include,but are not limited to: (1) aluminum salts (alum), such as aluminumhydroxide, aluminum phosphate, aluminum sulfate, etc; (2) oil-in-wateremulsion formulations (with or without other specific immunostimulatingagents such as muramyl peptides (see below) or bacterial cell wallcomponents), such as for example (a) MF59™ (WO 90/14837; Chapter 10 inVaccine design: the subunit and adjuvant approach, eds. Powell & Newman,Plenum Press 1995), containing 5% Squalene, 0.5% Tween 80, and 0.5% Span85 (optionally containing various amounts of MTP-PE (see below),although not required) formulated into submicron particles using amicrofluidizer such as Model 110Y microfluidizer (Microfluidics, Newton,Mass.), (b) SAF, containing 10% Squalane, 0.4% Tween 80, 5%pluronic-blocked polymer L121, and thr-MDP (see below) eithermicrofluidized into a submicron emulsion or vortexed to generate alarger particle size emulsion, and (c) Ribi™ adjuvant system (RAS),(Ribi Immunochem, Hamilton, Mont.) containing 2% Squalene, 0.2% Tween80, and one or more bacterial cell wall components from the groupconsisting of monophosphorylipid A (MPL), trehalose dimycolate (TDM),and cell wall skeleton (CWS), preferably MPL+CWS (Detox™); (3) saponinadjuvants, such as Stimulon™ (Cambridge Bioscience, Worcester, Mass.)may be used or particles generated therefrom such as ISCOMs(immunostimulating complexes); (4) Complete Freund's Adjuvant (CFA) andIncomplete Freund's Adjuvant (IFA); (5) cytokines, such as interleukins(eg. IL-1, IL-2, IL-4, IL-5, IL-6, IL-7, IL-12, etc.), interferons (eg.gamma interferon), macrophage colony stimulating factor (M-CSF), tumornecrosis factor (TNF), etc; and (6) other substances that act asimmunostimulating agents to enhance the effectiveness of thecomposition. Alum and MF59™ are preferred.

As mentioned above, muramyl peptides include, but are not limited to,N-acetyl-muramyl-L-threonyl-D-isoglutamine (thr-MDP),N-acetyl-normuramyl-L-alanyl-D-isoglutamine (nor-MDP),N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1′-2′-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamine(MTP-PE), etc.

The immunogenic compositions (eg. the immunisingantigen/immunogen/polypeptide/protein/nucleic acid, pharmaceuticallyacceptable carrier, and adjuvant) typically will contain diluents, suchas water, saline, glycerol, ethanol, etc. Additionally, auxiliarysubstances, such as wetting or emulsifying agents, pH bufferingsubstances, and the like, may be present in such vehicles.

Typically, the immunogenic compositions are prepared as injectables,either as liquid solutions or suspensions; solid forms suitable forsolution in, or suspension in, liquid vehicles prior to injection mayalso be prepared. The preparation also may be emulsified or encapsulatedin liposomes for enhanced adjuvant effect, as discussed above underpharmaceutically acceptable carriers.

Immunogenic compositions used as vaccines comprise an immunologicallyeffective amount of the antigenic or immunogenic polypeptides, as wellas any other of the above-mentioned components, as needed. By“immunologically effective amount”, it is meant that the administrationof that amount to an individual, either in a single dose or as part of aseries, is effective for treatment or prevention. This amount variesdepending upon the health and physical condition of the individual to betreated, the taxonomic group of individual to be treated (eg. nonhumanprimate, primate, etc.), the capacity of the individual's immune systemto synthesize antibodies, the degree of protection desired, theformulation of the vaccine, the treating doctor's assessment of themedical situation, and other relevant factors. It is expected that theamount will fall in a relatively broad range that can be determinedthrough routine trials.

The immunogenic compositions are conventionally administeredparenterally, eg. by injection, either subcutaneously, intramuscularly,or transdermally/transcutaneously (eg. WO98/20734). Additionalformulations suitable for other modes of administration include oral andpulmonary formulations, suppositories, and transdermal applications.Dosage treatment may be a single dose schedule or a multiple doseschedule. The vaccine may be administered in conjunction with otherimmunoregulatory agents.

As an alternative to protein-based vaccines, DNA vaccination may beemployed [eg. Robinson & Torres (1997) Seminars in Immunology 9:271-283;Donnelly et al. (1997) Annu Rev Immunol 15:617-648; see later herein].

Gene Delivery Vehicles

Gene therapy vehicles for delivery of constructs including a codingsequence of a therapeutic of the invention, to be delivered to themammal for expression in the mammal, can be administered either locallyor systemically. These constructs can utilize viral or non-viral vectorapproaches in in vivo or ex vivo modality. Expression of such codingsequence can be induced using endogenous mammalian or heterologouspromoters. Expression of the coding sequence in vivo can be eitherconstitutive or regulated.

The invention includes gene delivery vehicles capable of expressing thecontemplated nucleic acid sequences. The gene delivery vehicle ispreferably a viral vector and, more preferably, a retroviral,adenoviral, adeno-associated viral (AAV), herpes viral, or alphavirusvector. The viral vector can also be an astrovirus, coronavirus,orthomyxovirus, papovavirus, paramyxovirus, parvovirus, picornavirus,poxvirus, or togavirus viral vector. See generally, Jolly (1994) CancerGene Therapy 1:51-64; Kimura (1994) Human Gene Therapy 5:845-852;Connelly (1995) Human Gene Therapy 6:185-193; and Kaplitt (1994) NatureGenetics 6:148-153.

Retroviral vectors are well known in the art and we contemplate that anyretroviral gene therapy vector is employable in the invention, includingB, C and D type retroviruses, xenotropic retroviruses (for example,NZB-X1, NZB-X2 and NZB9-1 (see O'Neill (1985) J. Virol. 53:160)polytropic retroviruses eg. MCF and MCF-M LV (see Kelly (1983) J. Virol.45:291), spumaviruses and lentiviruses. See RNA Tumor Viruses, SecondEdition, Cold Spring Harbor Laboratory, 1985.

Portions of the retroviral gene therapy vector may be derived fromdifferent retroviruses. For example, retrovector LTRs may be derivedfrom a Murine Sarcoma Virus, a tRNA binding site from a Rous SarcomaVirus, a packaging signal from a Murine Leukemia Virus, and an origin ofsecond strand synthesis from an Avian Leukosis Virus.

These recombinant retroviral vectors may be used to generatetransduction competent retroviral vector particles by introducing theminto appropriate packaging cell lines (see U.S. Pat. No. 5,591,624).Retrovirus vectors can be constructed for site-specific integration intohost cell DNA by incorporation of a chimeric integrase enzyme into theretroviral particle (see WO96/37626). It is preferable that therecombinant viral vector is a replication defective recombinant virus.

Packaging cell lines suitable for use with the above-describedretrovirus vectors are well known in the art, are readily prepared (seeWO95/30763 and WO92/05266), and can be used to create producer celllines (also termed vector cell lines or “VCLs”) for the production ofrecombinant vector particles. Preferably, the packaging cell lines aremade from human parent cells (eg. HT1080 cells) or mink parent celllines, which eliminates inactivation in human serum.

Preferred retroviruses for the construction of retroviral gene therapyvectors include Avian Leukosis Virus, Bovine Leukemia, Virus, MurineLeukemia Virus, Mink-Cell Focus-Inducing Virus, Murine Sarcoma Virus,Reticuloendotheliosis Virus and Rous Sarcoma Virus. Particularlypreferred Murine Leukemia Viruses include 4070A and 1504A (Hartley andRowe (1976) J Virol 19:19-25), Abelson (ATCC No. VR-999), Friend (ATCCNo. VR-245), Graffi, Gross (ATCC Nol VR-590), Kirsten, Harvey SarcomaVirus and Rauscher (ATCC No. VR-998) and Moloney Murine Leukemia Virus(ATCC No. VR-190). Such retroviruses may be obtained from depositoriesor collections such as the American Type Culture Collection (“ATCC”) inRockville, Md. or isolated from known sources using commonly availabletechniques.

Exemplary known retroviral gene therapy vectors employable in thisinvention include those described in patent applications GB2200651,EP0415731, EP0345242, EP0334301, WO89/02468; WO89/05349, WO89/09271,WO90/02806, WO90/07936, WO94/03622, WO93/25698, WO93/25234, WO93/11230,WO93/10218, WO91/02805, WO91/02825, WO95/07994, U.S. Pat. No. 5,219,740,U.S. Pat. No. 4,405,712, U.S. Pat. No. 4,861,719, U.S. Pat. No.4,980,289, U.S. Pat. No. 4,777,127, U.S. Pat. No. 5,591,624. See alsoVile (1993) Cancer Res 53:3860-3864; Vile (1993) Cancer Res 53:962-967;Ram (1993) Cancer Res 53 (1993) 83-88; Takamiya (1992) J Neurosci Res33:493-503; Baba (1993) J Neurosurg 79:729-735; Mann (1983) Cell 33:153;Cane (1984) Proc Natl Acad Sci 81:6349; and Miller (1990) Human GeneTherapy 1.

Human adenoviral gene therapy vectors are also known in the art andemployable in this invention. See, for example, Berkner (1988)Biotechniques 6:616 and Rosenfeld (1991) Science 252:431, andWO93/07283, WO93/06223, and WO93/07282. Exemplary known adenoviral genetherapy vectors employable in this invention include those described inthe above referenced documents and in WO94/12649, WO93/03769,WO93/19191, WO94/28938, WO95/11984, WO95/00655, WO95/27071, WO95/29993,WO95/34671, WO96/05320, WO94/08026, WO94/11506, WO93/06223, WO94/24299,WO95/14102, WO95/24297, WO95/02697, WO94/28152, WO94/24299, WO95/09241,WO95/25807, WO95/05835, WO94/18922 and WO95/09654. Alternatively,administration of DNA linked to killed adenovirus as described in Curiel(1992) Hum. Gene Ther. 3:147-154 may be employed. The gene deliveryvehicles of the invention also include adenovirus associated virus (AAV)vectors. Leading and preferred examples of such vectors for use in thisinvention are the AAV-2 based vectors disclosed in Srivastava,WO93/09239. Most preferred AAV vectors comprise the two AAV invertedterminal repeats in which the native D-sequences are modified bysubstitution of nucleotides, such that at least 5 native nucleotides andup to 18 native nucleotides, preferably at least 10 native nucleotidesup to 18 native nucleotides, most preferably 10 native nucleotides areretained and the remaining nucleotides of the D-sequence are deleted orreplaced with non-native nucleotides. The native D-sequences of the AAVinverted terminal repeats are sequences of 20 consecutive nucleotides ineach AAV inverted terminal repeat (ie. there is one sequence at eachend) which are not involved in HP formation. The non-native replacementnucleotide may be any nucleotide other than the nucleotide found in thenative D-sequence in the same position. Other employable exemplary AAVvectors are pWP-19, pWN-1, both of which are disclosed in Nahreini(1993) Gene 124:257-262. Another example of such an AAV vector ispsub201 (see Samulski (1987) J. Virol. 61:3096). Another exemplary AAVvector is the Double-D ITR vector. Construction of the Double-D ITRvector is disclosed in U.S. Pat. No. 5,478,745. Still other vectors arethose disclosed in Carter U.S. Pat. No. 4,797,368 and Muzyczka U.S. Pat.No. 5,139,941, Chartejee U.S. Pat. No. 5,474,935, and Kotin WO94/288157.Yet a further example of an AAV vector employable in this invention isSSV9AFABTKneo, which contains the AFP enhancer and albumin promoter anddirects expression predominantly in the liver. Its structure andconstruction are disclosed in Su (1996) Human Gene Therapy 7:463-470.Additional AAV gene therapy vectors are described in U.S. Pat. No.5,354,678, U.S. Pat. No. 5,173,414, U.S. Pat. No. 5,139,941, and U.S.Pat. No. 5,252,479.

The gene therapy vectors of the invention also include herpes vectors.Leading and preferred examples are herpes simplex virus vectorscontaining a sequence encoding a thymidine kinase polypeptide such asthose disclosed in U.S. Pat. No. 5,288,641 and EP0176170 (Roizman).Additional exemplary herpes simplex virus vectors include HFEWICP6-LacZdisclosed in WO95/04139 (Wistar Institute), pHSVlac described in Geller(1988) Science 241:1667-1669 and in WO90/09441 and WO92/07945, HSVUs3::pgC-lacZ described in Fink (1992) Human Gene Therapy 3:11-19 andHSV 7134, 2 RH 105 and GAL4 described in EP 0453242 (Breakefield), andthose deposited with the ATCC as accession numbers ATCC VR-977 and ATCCVR-260.

Also contemplated are alpha virus gene therapy vectors that can beemployed in this invention. Preferred alpha virus vectors are Sindbisviruses vectors. Togaviruses, Semliki Forest virus (ATCC VR-67; ATCCVR-1247), Middleberg virus (ATCC VR-370), Ross River virus (ATCC VR-373;ATCC VR-1246), Venezuelan equine encephalitis virus (ATCC VR923; ATCCVR-1250; ATCC VR-1249; ATCC VR-532), and those described in U.S. Pat.Nos. 5,091,309, 5,217,879, and WO92/10578. More particularly, thosealpha virus vectors described in U.S. Ser. No. 08/405,627, filed Mar.15, 1995, WO94/21792, WO92/10578, WO95/07994, U.S. Pat. No. 5,091,309and U.S. Pat. No. 5,217,879 are employable. Such alpha viruses may beobtained from depositories or collections such as the ATCC in Rockville,Md. or isolated from known sources using commonly available techniques.Preferably, alphavirus vectors with reduced cytotoxicity are used (seeU.S. Ser. No. 08/679,640).

DNA vector systems such as eukarytic layered expression systems are alsouseful for expressing the nucleic acids of the invention. See WO95/07994for a detailed description of eukaryotic layered expression systems.Preferably, the eukaryotic layered expression systems of the inventionare derived from alphavirus vectors and most preferably from Sindbisviral vectors.

Other viral vectors suitable for use in the present invention includethose derived from poliovirus, for example ATCC VR-58 and thosedescribed in Evans, Nature 339 (1989)385 and Sabin (1973) J. Biol.Standardization 1:115; rhinovirus, for example ATCC VR-1110 and thosedescribed in Arnold (1990) J Cell Biochem L401; pox viruses such ascanary pox virus or vaccinia virus, for example ATCC VR-111 and ATCCVR-2010 and those described in Fisher-Hoch (1989) Proc Nan Acad Sci86:317; Flexner (1989) Ann NY Acad Sci 569:86, Flexner (1990) Vaccine8:17; in U.S. Pat. No. 4,603,112 and U.S. Pat. No. 4,769,330 andWO89/01973; SV40 virus, for example ATCC VR-305 and those described inMulligan (1979) Nature 277:108 and Madzak (1992) J Gen Virol 73:1533;influenza virus, for example ATCC VR-797 and recombinant influenzaviruses made employing reverse genetics techniques as described in U.S.Pat. No. 5,166,057 and in Enami (1990) Proc Nall Acad Sci 87:3802-3805;Enami & Palese (1991) J Virol 65:2711-2713 and Luytjes (1989) Cell59:110, (see also McMichael (1983) NEJ Med 309:13, and Yap (1978) Nature273:238 and Nature (1979) 277:108); human immunodeficiency virus asdescribed in EP-0386882 and in Buchschacher (1992) J. Virol. 66:2731;measles virus, for example ATCC VR-67 and VR-1247 and those described inEP-0440219; Aura virus, for example ATCC VR-368; Bebaru virus, forexample ATCC VR-600 and ATCC VR-1240; Cabassou virus, for example ATCCVR-922; Chikungunya virus, for example ATCC VR-64 and ATCC VR-1241; FortMorgan Virus, for example ATCC VR-924; Getah virus, for example ATCCVR-369 and ATCC VR-1243; Kyzylagach virus, for example ATCC VR-927;Mayaro virus, for example ATCC VR-66; Mucambo virus, for example ATCCVR-580 and ATCC VR-1244; Ndumu virus, for example ATCC VR-371; Pixunavirus, for example ATCC VR-372 and ATCC VR-1245; Tonate virus, forexample ATCC VR-925; Triniti virus, for example ATCC VR-469; Una virus,for example ATCC VR-374; Whataroa virus, for example ATCC VR-926;Y-62-33 virus, for example ATCC VR-375; O'Nyong virus, Easternencephalitis virus, for example ATCC VR-65 and ATCC VR-1242; Westernencephalitis virus, for example ATCC VR-70, ATCC VR-1251, ATCC VR-622and ATCC VR-1252; and coronavirus, for example ATCC VR-740 and thosedescribed in Hamre (1966) Proc Soc Exp Biol Med 121:190.

Delivery of the compositions of this invention into cells is not limitedto the above mentioned viral vectors. Other delivery methods and mediamay be employed such as, for example, nucleic acid expression vectors,polycationic condensed DNA linked or unlinked to killed adenovirusalone, for example see U.S. Ser. No. 08/366,787, filed Dec. 30, 1994 andCurie] (1992) Hum Gene Titer 3:147-154 ligand linked DNA, for examplesee Wu (1989) J Biol Chem 264:16985-16987, eucaryotic cell deliveryvehicles cells, for example see U.S. Ser. No. 08/240,030, filed May 9,1994, and U.S. Ser. No. 08/404,796, deposition of photopolymerizedhydrogel materials, hand-held gene transfer particle gun, as describedin U.S. Pat. No. 5,149,655, ionizing radiation as described in U.S. Pat.No. 5,206,152 and in WO92/11033, nucleic charge neutralization or fusionwith cell membranes. Additional approaches are described in Philip(1994) Mol Cell Biol 14:2411-2418 and in Woffendin (1994) Proc Natl AcadSci 91:1581-1585.

Particle mediated gene transfer may be employed, for example see U.S.Ser. No. 60/023,867. Briefly, the sequence can be inserted intoconventional vectors that contain conventional control sequences forhigh level expression, and then incubated with synthetic gene transfermolecules such as polymeric DNA-binding cations like polylysine,protamine, and albumin, linked to cell targeting ligands such asasialoorosomucoid, as described in Wu & Wu (1987) J. Biol. Chem.262:4429-4432, insulin as described in Hucked (1990) Biochem Pharmacol40:253-263, galactose as described in Plank (1992) Bioconjugate Chem3:533-539, lactose or transferrin.

Naked DNA may also be employed. Exemplary naked DNA introduction methodsare described in WO 90/11092 and U.S. Pat. No. 5,580,859. Uptakeefficiency may be improved using biodegradable latex beads. DNA coatedlatex beads are efficiently transported into cells after endocytosisinitiation by the beads. The method may be improved further by treatmentof the beads to increase hydrophobicity and thereby facilitatedisruption of the endosome and release of the DNA into the cytoplasm.

Liposomes that can act as gene delivery vehicles are described in U.S.Pat. No. 5,422,120, WO95/13796, WO94/23697, WO91/14445 and EP-524,968.As described in U.S. Ser. No. 60/023,867, on non-viral delivery, thenucleic acid sequences encoding a polypeptide can be inserted intoconventional vectors that contain conventional control sequences forhigh level expression, and then be incubated with synthetic genetransfer molecules such as polymeric DNA-binding cations likepolylysine, protamine, and albumin, linked to cell targeting ligandssuch as asialoorosomucoid, insulin, galactose, lactose, or transferrin.Other delivery systems include the use of liposomes to encapsulate DNAcomprising the gene under the control of a variety of tissue-specific orubiquitously-active promoters. Further non-viral delivery suitable foruse includes mechanical delivery systems such as the approach describedin Woffendin et al (1994) Proc. Natl. Acad. Sci. USA 91(24):11581-11585.Moreover, the coding sequence and the product of expression of such canbe delivered through deposition of photopolymerized hydrogel materials.Other conventional methods for gene delivery that can be used fordelivery of the coding sequence include, for example, use of hand-heldgene transfer particle gun, as described in U.S. Pat. No. 5,149,655; useof ionizing radiation for activating transferred gene, as described inU.S. Pat. No. 5,206,152 and WO92/11033

Exemplary liposome and polycationic gene delivery vehicles are thosedescribed in U.S. Pat. Nos. 5,422,120 and 4,762,915; in WO 95/13796;WO94/23697; and WO91/14445; in EP-0524968; and in Stryer, Biochemistry,pages 236-240 (1975) W.H. Freeman, San Francisco; Szoka (1980) BiochemBiophys Acta 600:1; Bayer (1979) Biochem Biophys Acta 550:464; Rivnay(1987) Meth Enzymol 149:119; Wang (1987) Proc Natl Acad Sci 84:7851;Plant (1989) Anal Biochem 176:420.

A polynucleotide composition can comprises therapeutically effectiveamount of a gene therapy vehicle, as the term is defined above. Forpurposes of the present invention, an effective dose will be from about0.01 mg/kg to 50 mg/kg or 0.05 mg/kg to about 10 mg/kg of the DNAconstructs in the individual to which it is administered.

Delivery Methods

Once formulated, the polynucleotide compositions of the invention can beadministered (1) directly to the subject; (2) delivered ex vivo, tocells derived from the subject; or (3) in vitro for expression ofrecombinant proteins. The subjects to be treated can be mammals orbirds. Also, human subjects can be treated.

Direct delivery of the compositions will generally be accomplished byinjection, either subcutaneously, intraperitoneally, intravenously orintramuscularly or delivered to the interstitial space of a tissue. Thecompositions can also be administered into a lesion. Other modes ofadministration include oral and pulmonary administration, suppositories,and transdermal or transcutaneous applications (eg. see WO98/20734),needles, and gene guns or hyposprays. Dosage treatment may be a singledose schedule or a multiple dose schedule.

Methods for the ex vivo delivery and reimplantation of transformed cellsinto a subject are known in the art and described in eg. WO93/14778.Examples of cells useful in ex vivo applications include, for example,stem cells, particularly hematopoetic, lymph cells, macrophages,dendritic cells, or tumor cells.

Generally, delivery of nucleic acids for both ex vivo and in vitroapplications can be accomplished by the following procedures, forexample, dextran-mediated transfection, calcium phosphate precipitation,polybrene mediated transfection, protoplast fusion, electroporation,encapsulation of the polynucleotide(s) in liposomes, and directmicroinjection of the DNA into nuclei, all well known in the art.

Polynucleotide and Polypeptide Pharmaceutical Compositions

In addition to the pharmaceutically acceptable carriers and saltsdescribed above, the following additional agents can be used withpolynucleotide and/or polypeptide compositions.

A. Polypeptides

One example are polypeptides which include, without limitation:asioloorosomucoid (ASOR); transferrin; asialoglycoproteins; antibodies;antibody fragments; ferritin; interleukins; interferons, granulocyte,macrophage colony stimulating factor (GM-CSF), granulocyte colonystimulating factor (G-CSF), macrophage colony stimulating factor(M-CSF), stem cell factor and erythropoietin. Viral antigens, such asenvelope proteins, can also be used. Also, proteins from other invasiveorganisms, such as the 17 amino acid peptide from the circumsporozoiteprotein of plasmodium falciparum known as RH.

B. Hormones, Vitamins, etc.

Other groups that can be included are, for example: hormones, steroids,androgens, estrogens, thyroid hormone, or vitamins, folic acid.

C. Polyalkylenes, Polysaccharides, etc.

Also, polyalkylene glycol can be included with the desiredpolynucleotides/polypeptides. In a preferred embodiment, thepolyalkylene glycol is polyethlylene glycol. Iri addition, mono-, di-,or polysaccarides can be included. In a preferred embodiment of thisaspect, the polysaccharide is dextran or DEAE-dextran. Also, chitosanand poly(lactide-co-glycolide)

D. Lipids, and Liposomes

The desired polynucleotide/polypeptide can also be encapsulated inlipids or packaged in liposomes prior to delivery to the subject or tocells derived therefrom.

Lipid encapsulation is generally accomplished using liposomes which areable to stably bind or entrap and retain nucleic acid. The ratio ofcondensed polynucleotide to lipid preparation can vary but willgenerally be around 1:1 (mg DNA:micromoles lipid), or more of lipid. Fora review of the use of liposomes as carriers for delivery of nucleicacids, see, Hug and Sleight (1991) Biochim. Biophys. Acta. 1097:1-17;Straubinger (1983) Meth. Enzymol. 101:512-527.

Liposomal preparations for use in the present invention include cationic(positively charged), anionic (negatively charged) and neutralpreparations. Cationic liposomes have been shown to mediateintracellular delivery of plasmid DNA (Feigner (1987) Proc. Natl. Acad.Sci. USA 84:7413-7416); mRNA (Malone (1989) Proc. Natl. Acad. Sci. USA86:6077-6081); and purified transcription factors (Debs (1990) J. Biol.Chem. 265:10189-10192), in functional form.

Cationic liposomes are readily available. For example,N[1-2,3-dioleyloxy)propyl]-N,N,N-triethylammonium (DOTMA) liposomes areavailable under the trademark Lipofectin, from GIBCO BRL, Grand Island,N.Y. (See, also, Feigner supra). Other commercially available liposomesinclude transfectace (DDAB/DOPE) and DOTAP/DOPE (Boerhinger). Othercationic liposomes can be prepared from readily available materialsusing techniques well known in the art. See, eg. Szoka (1978) Proc.Natl. Acad. Sci. USA 75:4194-4198; WO90/11092 fora description of thesynthesis of DOTAP (1,2-bis(oleoyloxy)-3-(trimethylammonio)propane)liposomes.

Similarly, anionic and neutral liposomes are readily available, such asfrom Avanti Polar Lipids (Birmingham, Ala.), or can be easily preparedusing readily available materials. Such materials include phosphatidylcholine, cholesterol, phosphatidyl ethanolamine, dioleoylphosphatidylcholine (DOPC), dioleoylphosphatidyl glycerol (DOPG),dioleoylphoshatidyl ethanolamine (DOPE), among others. These materialscan also be mixed with the DOTMA and DOTAP starting materials inappropriate ratios. Methods for making liposomes using these materialsare well known in the art.

The liposomes can comprise multilammelar vesicles (MLVs), smallunilamellar vesicles (SUVs), or large unilamellar vesicles (LUVs). Thevarious liposome-nucleic acid complexes are prepared using methods knownin the art. See eg. Straubinger (1983) Meth. Immunol. 101:512-527; Szoka(1978) Proc. Natl. Acad. Sci. USA 75:4194-4198; Papahadjopoulos (1975)Biochim. Biophys. Acta 394:483; Wilson (1979) Cell 17:77); Deamer &Bangham (1976) Biochim. Biophys. Ada 443:629; Ostro (1977) Biochem.Biophys. Res. Commun. 76:836; Fraley (1979) Proc. Natl. Acad. Sci. USA76:3348); Enoch & Strittmatter (1979) Proc. Natl. Acad. Sci. USA 76:145;Fraley (1980) J. Biol. Chem. (1980) 255:10431; Szoka & Papahadjopoulos(1978) Proc. Natl. Acad. Sci. USA 75:145; and Schaefer-Ridder (1982)Science 215:166.

E. Lipoproteins

In addition, lipoproteins can be included with thepolynucleotide/polypeptide to be delivered. Examples of lipoproteins tobe utilized include: chylomicrons, HDL, IDL, LDL, and VLDL. Mutants,fragments, or fusions of these proteins can also be used. Also,modifications of naturally occurring lipoproteins can be used, such asacetylated LDL. These lipoproteins can target the delivery ofpolynucleotides to cells expressing lipoprotein receptors. Preferably,if lipoproteins are including with the polynucleotide to be delivered,no other targeting ligand is included in the composition.

Naturally occurring lipoproteins comprise a lipid and a protein portion.The protein portion are known as apoproteins. At the present,apoproteins A, B, C, D, and E have been isolated and identified. Atleast two of these contain several proteins, designated by Romannumerals, AI, AII, AIV; CI, CII, CIII.

A lipoprotein can comprise more than one apoprotein. For example,naturally occurring chylomicrons comprises of A, B, C, and E, over timethese lipoproteins lose A and acquire C and E apoproteins. VLDLcomprises A, B, C, and E apoproteins, LDL comprises apoprotein B; andHDL comprises apoproteins A, C, and E.

The amino acid of these apoproteins are known and are described in, forexample, Breslow (1985) Annu Rev. Biochem 54:699; Law (1986) Adv. ExpMed. Biol. 151:162; Chen (1986) J Biol Chem 261:12918; Kane (1980) ProcNatl Acad Sci USA 77:2465; and Utermann (1984) Hum Genet 65:232.

Lipoproteins contain a variety of lipids including, triglycerides,cholesterol (free and esters), and phopholipids. The composition of thelipids varies in naturally occurring lipoproteins. For example,chylomicrons comprise mainly triglycerides. A more detailed descriptionof the lipid content of naturally occurring lipoproteins can be found,for example, in Meth. Enzymol. 128 (1986). The composition of the lipidsare chosen to aid in conformation of the apoprotein for receptor bindingactivity. The composition of lipids can also be chosen to facilitatehydrophobic interaction and association with the polynucleotide bindingmolecule.

Naturally occurring lipoproteins can be isolated from serum byultracentrifugation, for instance. Such methods are described in Meth.Enzymol. (supra); Pitas (1980) J. Biochem. 255:5454-5460 and Mahey(1979) J. Clin. Invest 64:743-750. Lipoproteins can also be produced byin vitro or recombinant methods by expression of the apoprotein genes ina desired host cell. See, for example, Atkinson (1986) Annu Rev BiophysChem 15:403 and Radding (1958) Biochim Biophys Acta 30: 443.Lipoproteins can also be purchased from commercial suppliers, such asBiomedical Techniologies, Inc., Stoughton, Mass., USA. Furtherdescription of lipoproteins can be found in Zuckermann et al.PCT/US97/14465.

F. Polycationic Agents

Polycationic agents can be included, with or without lipoprotein, in acomposition with the desired polynucleotide/polypeptide to be delivered.

Polycationic agents, typically, exhibit a net positive charge atphysiological relevant pH and are capable of neutralizing the electricalcharge of nucleic acids to facilitate delivery to a desired location.These agents have both in vitro, ex vivo, and in vivo applications.Polycationic agents can be used to deliver nucleic acids to a livingsubject either intramuscularly, subcutaneously, etc.

The following are examples of useful polypeptides as polycationicagents: polylysine, polyarginine, polyornithine, and protamine. Otherexamples include histones, protamines, human serum albumin, DNA bindingproteins, non-histone chromosomal proteins, coat proteins from DNAviruses, such as (X174, transcriptional factors also contain domainsthat bind DNA and therefore may be useful as nucleic aid condensingagents. Briefly, transcriptional factors such as C/CEBP, c-jun, c-fos,AP-1, AP-2, AP-3, CPF, Prot-1, Sp-1, Oct-1, Oct-2, CREP, and TFIIDcontain basic domains that bind DNA sequences.

Organic polycationic agents include: spermine, spermidine, andpurtrescine.

The dimensions and of the physical properties of a polycationic agentcan be extrapolated from the list above, to construct other polypeptidepolycationic agents or to produce synthetic polycationic agents.

Synthetic polycationic agents which are useful include, for example,DEAE-dextran, polybrene. Lipofectin™, and lipofectAMINE™ are monomersthat form polycationic complexes when combined withpolynucleotides/polypeptides.

Immunodiagnostic Assays

Neisserial antigens of the invention can be used in immunoassays todetect antibody levels (or, conversely, anti-Neisserial antibodies canbe used to detect antigen levels). Immunoassays based on well defined,recombinant antigens can be developed to replace invasive diagnosticsmethods. Antibodies to Neisserial proteins within biological samples,including for example, blood or serum samples, can be detected. Designof the immunoassays is subject to a great deal of variation, and avariety of these are known in the art. Protocols for the immunoassay maybe based, for example, upon competition, or direct reaction, or sandwichtype assays. Protocols may also, for example, use solid supports, or maybe by immunoprecipitation. Most assays involve the use of labeledantibody or polypeptide; the labels may be, for example, fluorescent,chemiluminescent, radioactive, or dye molecules. Assays which amplifythe signals from the probe are also known; examples of which are assayswhich utilize biotin and avidin, and enzyme-labeled and mediatedimmunoassays, such as ELISA assays.

Kits suitable for immunodiagnosis and containing the appropriate labeledreagents are constructed by packaging the appropriate materials,including the compositions of the invention, in suitable containers,along with the remaining reagents and materials (for example, suitablebuffers, salt solutions, etc.) required for the conduct of the assay, aswell as suitable set of assay instructions.

Nucleic Acid Hybridisation

“Hybridization” refers to the association of two nucleic acid sequencesto one another by hydrogen bonding. Typically, one sequence will befixed to a solid support and the other will be free in solution. Then,the two sequences will be placed in contact with one another underconditions that favor hydrogen bonding. Factors that affect this bondinginclude: the type and volume of solvent; reaction temperature; time ofhybridization; agitation; agents to block the non-specific attachment ofthe liquid phase sequence to the solid support (Denhardt's reagent orBLOTTO); concentration of the sequences; use of compounds to increasethe rate of association of sequences (dextran sulfate or polyethyleneglycol); and the stringency of the washing conditions followinghybridization. See Sambrook et al. [supra] Volume 2, chapter 9, pages9.47 to 9.57.

“Stringency” refers to conditions in a hybridization reaction that favorassociation of very similar sequences over sequences that differ. Forexample, the combination of temperature and salt concentration should bechosen that is approximately 120 to 200° C. below the calculated Tm ofthe hybrid under study. The temperature and salt conditions can often bedetermined empirically in preliminary experiments in which samples ofgenomic DNA immobilized on filters are hybridized to the sequence ofinterest and then washed under conditions of different stringencies. SeeSambrook et al. at page 9.50.

Variables to consider when performing, for example, a Southern blot are(1) the complexity of the DNA being blotted and (2) the homology betweenthe probe and the sequences being detected. The total amount of thefragment(s) to be studied can vary a magnitude of 10, from 0.1 to 1 μgfor a plasmid or phage digest to 10⁻⁹ to 10⁻⁸ g for a single copy genein a highly complex eukaryotic genome. For lower complexitypolynucleotides, substantially shorter blotting, hybridization, andexposure times, a smaller amount of starting polynucleotides, and lowerspecific activity of probes can be used. For example, a single-copyyeast gene can be detected with an exposure time of only 1 hour startingwith 1 μg of yeast DNA, blotting for two hours, and hybridizing for 4-8hours with a probe of 10⁸ cpm/μg. For a single-copy mammalian gene aconservative approach would start with 10 μg of DNA, blot overnight, andhybridize overnight in the presence of 10% dextran sulfate using a probeof greater than 10⁸ cpm/μg, resulting in an exposure time of ˜24 hours.

Several factors can affect the melting temperature (Tm) of a DNA-DNAhybrid between the probe and the fragment of interest, and consequently,the appropriate conditions for hybridization and washing. In many casesthe probe is not 100% homologous to the fragment. Other commonlyencountered variables include the length and total G+C content of thehybridizing sequences and the ionic strength and formamide content ofthe hybridization buffer. The effects of all of these factors can beapproximated by a single equation:

Tm=81+16.6(log₁₀Ci)+0.4[%(G+C)]−0.6(% formamide)−600/n−1.5(% mismatch).

where Ci is the salt concentration (monovalent ions) and n is the lengthof the hybrid in base pairs (slightly modified from Meinkoth & Wahl(1984) Anal. Biochem. 138: 267-284).

In designing a hybridization experiment, some factors affecting nucleicacid hybridization can be conveniently altered. The temperature of thehybridization and washes and the salt concentration during the washesare the simplest to adjust. As the temperature of the hybridizationincreases (ie. stringency), it becomes less likely for hybridization tooccur between strands that are nonhomologous, and as a result,background decreases. If the radiolabeled probe is not completelyhomologous with the immobilized fragment (as is frequently the case ingene family and interspecies hybridization experiments), thehybridization temperature must be reduced, and background will increase.The temperature of the washes affects the intensity of the hybridizingband and the degree of background in a similar manner. The stringency ofthe washes is also increased with decreasing salt concentrations.

In general, convenient hybridization temperatures in the presence of 50%formamide are 42° C. for a probe with is 95% to 100% homologous to thetarget fragment, 37° C. for 90% to 95% homology, and 32° C. for 85% to90% homology. For lower homologies, formamide content should be loweredand temperature adjusted accordingly, using the equation above. If thehomology between the probe and the target fragment are not known, thesimplest approach is to start with both hybridization and washconditions which are nonstringent. If non-specific bands or highbackground are observed after autoradiography, the filter can be washedat high stringency and reexposed. If the time required for exposuremakes this approach impractical, several hybridization and/or washingstringencies should be tested in parallel.

Nucleic Acid Probe Assays

Methods such as PCR, branched DNA probe assays, or blotting techniquesutilizing nucleic acid probes according to the invention can determinethe presence of cDNA or mRNA. A probe is said to “hybridize” with asequence of the invention if it can form a duplex or double strandedcomplex, which is stable enough to be detected.

The nucleic acid probes will hybridize to the Neisserial nucleotidesequences of the invention (including both sense and antisense strands).Though many different nucleotide sequences will encode the amino acidsequence, the native Neisserial sequence is preferred because it is theactual sequence present in cells. mRNA represents a coding sequence andso a probe should be complementary to the coding sequence;single-stranded cDNA is complementary to mRNA, and so a cDNA probeshould be complementary to the non-coding sequence.

The probe sequence need not be identical to the Neisserial sequence (orits complement)—some variation in the sequence and length can lead toincreased assay sensitivity if the nucleic acid probe can form a duplexwith target nucleotides, which can be detected. Also, the nucleic acidprobe can include additional nucleotides to stabilize the formed duplex.Additional Neisserial sequence may also be helpful as a label to detectthe formed duplex. For example, a non-complementary nucleotide sequencemay be attached to the 5′ end of the probe, with the remainder of theprobe sequence being complementary to a Neisserial sequence.Alternatively, non-complementary bases or longer sequences can beinterspersed into the probe, provided that the probe sequence hassufficient complementarity with the a Neisserial sequence in order tohybridize therewith and thereby form a duplex which can be detected.

The exact length and sequence of the probe will depend on thehybridization conditions, such as temperature, salt condition and thelike. For example, for diagnostic applications, depending on thecomplexity of the analyte sequence, the nucleic acid probe typicallycontains at least 10-20 nucleotides, preferably 15-25, and morepreferably at least 30 nucleotides, although it may be shorter thanthis. Short primers generally require cooler temperatures to formsufficiently stable hybrid complexes with the template.

Probes may be produced by synthetic procedures, such as the triestermethod of Matteucci et al. [J. Am. Chem. Soc. (1981) 103:3185], oraccording to Urdea et al. [Proc. Natl. Acad. Sci. USA (1983) 80: 7461],or using commercially available automated oligonucleotide synthesizers.

The chemical nature of the probe can be selected according topreference. For certain applications, DNA or RNA are appropriate. Forother applications, modifications may be incorporated eg. backbonemodifications, such as phosphorothioates or methylphosphonates, can beused to increase in vivo half-life, alter RNA affinity, increasenuclease resistance etc. [eg. see Agrawal & Iyer (1995) Curr OpinBiotechnol 6:12-19; Agrawal (1996) TIBTECH 14:376-387]; analogues suchas peptide nucleic acids may also be used [eg. see Corey (1997) TIBTECH15:224-229; Buchardt et al. (1993) TIBTECH 11:384-386].

Alternatively, the polymerase chain reaction (PCR) is another well-knownmeans for detecting small amounts of target nucleic acids. The assay isdescribed in: Mullis et al. [Meth. Enzymol. (1987) 155: 335-350]; U.S.Pat. Nos. 4,683,195 and 4,683,202. Two “primer” nucleotides hybridizewith the target nucleic acids and are used to prime the reaction. Theprimers can comprise sequence that does not hybridize to the sequence ofthe amplification target (or its complement) to aid with duplexstability or, for example, to incorporate a convenient restriction site.Typically, such sequence will flank the desired Neisserial sequence.

A thermostable polymerase creates copies of target nucleic acids fromthe primers using the original target nucleic acids as a template. Aftera threshold amount of target nucleic acids are generated by thepolymerase, they can be detected by more traditional methods, such asSouthern blots. When using the Southern blot method, the labelled probewill hybridize to the Neisserial sequence (or its complement).

Also, mRNA or cDNA can be detected by traditional blotting techniquesdescribed in Sambrook et al [supra]. mRNA, or cDNA generated from mRNAusing a polymerase enzyme, can be purified and separated using gelelectrophoresis. The nucleic acids on the gel are then blotted onto asolid support, such as nitrocellulose. The solid support is exposed to alabelled probe and then washed to remove any unhybridized probe. Next,the duplexes containing the labeled probe are detected. Typically, theprobe is labelled with a radioactive moiety.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A-E: For ORF37-1, (A) shows the results of affinity purificationof the GST-fusion protein, (B) shows the results of expression of theHis-fusion in E. coli. Purified GST-fusion protein was used to immunisemice, whose sera were used for ELISA (positive result), (C) shows FACSanalysis, and (D) shows a bactericidal assay (FIG. 1D), and (E) showsplots of hydrophilicity, antigenic index, and AMPHI regions for ORF37-1.

FIG. 2A-B: For ORF5-1, (A) shows the results of affinity purification ofthe GST-fusion protein, and (B) shows the Western blot analysis of serafrom mice immunized with purified GST-fusion protein.

FIG. 3A-D: For ORF2-1, (A) shows the results of affinity purification ofthe GST-fusion protein, (B) shows the results of expression of theHis-fusion in E. coli, (C) shows the Western blot analysis of sera frommice immunized with turified GST-fusion protein, (D) shows the ELISA(positive result), and (D) shows the FACS analysis.

FIG. 4A-C: For ORF15-1, (A) shows the results of affinity purificationof the GST-fusion protein, (B) shows the results of expression of theHis-fusion in E. coli, and (C) shows the Western blot analysis of serafrom mice immunized with purified GST-fusion protein.

FIG. 5A-C: For ORF22-1, (A) shows the results of affinity purificationof the GST-fusion protein, (B) shows the results of expression of theHis-fusion in E. coli, and (C) shows the FACS analysis using sera frommice immunized with the purified GST-fusion protein.

FIG. 6A-B: For ORF28-1, (A) shows the results of affinity purificationof the GST-fusion protein, and (B) shows the results of expression ofthe His-fusion in E. coli.

FIG. 7A-B: For ORF32-1, (A) shows the results of affinity purificationof the His-fusion protein, and (B) shows the results of expression ofthe GST-fusion in E. coli.

FIG. 8A-F: For ORF4-1, (A) shows the results of affinity purification ofthe His-fusion, (B) shows the results of affinity purification of theGST-fusion proteins, (C) shows the Western blot analysis of sera frommice immunized with the His-fusion protein, (D) shows the FACS analysis,(E) shows a bactericidal assay, and (F) shows plots of hydrophilicity,antigenic index, and AMPHI regions for ORF37-1.

FIG. 9 shows plots of hydrophilicity, antigenic index, and AMPHI regionsfor ORF61-1.

FIG. 10A-C: For ORF76-1, (A) shows the results of affinity purificationof the His-fusion protein, (B) shows the Western blot analysis of serafrom mice immunized with the purified His-fusion protein, and (C) showsthe FACS analysis.

FIG. 11 shows the results of affinity purification of the GST-ORF89-1fusion protein.

FIG. 12A-E: For ORF97-1, (A) show the results of affinity purificationof the GST-fusion protein, (B) shows the results of affinitypurification of the His-fusion protein, (C) shows the Western blotanalysis of sera from mice immunized with purified GST-fusion protein,(D) shows the FACS analysis, and (E) shows plots of hydrophilicity,antigenic index, and AMPHI regions for ORF97-1.

FIG. 13A-C: For ORF106-1, (A) shows the results of affinity purificationof the His-fusion protein, (B) shows the results of expression of theGST-fusion in E. coli, (C) shows the FACS analysis of sera from miceimmunized with the purified His-fusion protein.

FIG. 14A-B: For ORF138-1, (A) shows the results of affinity purificationof the GST-fusion protein, and (B) shows the FACS analysis of sera frommice immunized with the purified GST-fusion protein.

FIG. 15A-C: For ORF23-1, (A) shows the results of affinity purificationof the His-fusion protein, (B) shows the results of expression of theGST-fusion in E. coli, (C) shows the Western blot analysis of sera frommice immunized with the purified His-fusion protein.

FIG. 16A-E: For ORF25-1, (A) shows the results of affinity purificationof the GST-fusion protein, (B) shows the results of expression of theHis-fusion in E. coli, (C) shows the Western blot analysis of sera frommice immunized with purified His-fusion protein, (D) shows the FACSanalysis, and (E) shows plots of hydrophilicity, antigenic index, andAMPHI regions for ORF25-1.

FIG. 17A-B: For ORF27-1, (A) shows the results of affinity purificationof the GST-fusion protein, and (B) shows the results of expression ofthe His-fusion in E. coli.

FIG. 18A-B: For ORF79-1, (A) shows the results of affinity purificationof the His-fusion protein, and (B) shows the FACS analysis of sera frommice immunized with purified His-fusion protein.

FIG. 19A-D: For ORF85a, (A) shows the results of affinity purificationof the GST-fusion protein, (B) shows Western blot analysis of sera frommice immunized with purified GST-fusion protein, (C) shows FACSanalysis, and (D) shows plots of hydrophilicity, antigenic index, andAMPHI regions for ORF85a.

FIG. 20A-C: For ORF132-1, (A) shows the results of affinity purificationof the His-fusion protein, (B) shows the results of expression of theGST-fusion in E. coli, (C) shows the FACS analysis of sera from miceimmunized with purified His-fusion protein.

EXAMPLES

The examples describe nucleic acid sequences which have been identifiedin N. meningitidis, along with their putative translation products, andalso those of N. gonorrhoeae. Not all of the nucleic acid sequences arecomplete i.e. they encode less than the full-length wild-type protein.

The examples are generally in the following format:

-   -   a nucleotide sequence which has been identified in N.        meningitidis (strain B)    -   the putative translation product of this sequence    -   a computer analysis of the translation product based on database        comparisons    -   corresponding gene and protein sequences identified in N.        meningitidis (strain A) and in N. gonorrhoeae    -   a description of the characteristics of the proteins which        indicates that they might be suitably antigenic    -   results of biochemical analysis (expression, purification,        ELISA, FACS etc.)    -   The examples typically include details of sequence identity        between species and strains. Proteins that are similar in        sequence are generally similar in both structure and function,        and the sequence identity often indicates a common evolutionary        origin. Comparison with sequences of proteins of known function        is widely used as a guide for the assignment of putative protein        function to a new sequence and has proved particularly useful in        whole-genome analyses.

Sequence comparisons were performed at NCBI(http://www.ncbi.nlm.nih.gov) using the algorithms BLAST, BLAST2,BLASTn, BLASTp, tBLASTn, BLASTx, & tBLASTx [eg. see also Altschul et al.(1997) Gapped BLAST and PSI-BLAST: a new generation of protein databasesearch programs. Nucleic Acids Research 25:2289-3402]. Searches wereperformed against the following databases: non-redundantGenBank+EMBL+DDBJ+PDB sequences and non-redundant GenBank CDStranslations+PDB+SwissProt+SPupdate+PIR sequences.

To compare Meningococcal and Gonococcal sequences, the tBLASTx algorithmwas used, as implemented at http://www.genome.ou.edu/gono_blast.html.The FASTA algorithm was also used to compare the ORFs (from GCGWisconsin Package, version 9.0).

Dots within nucleotide sequences (eg. position 495 in SEQ ID 11)represent nucleotides which have been arbitrarily introduced in order tomaintain a reading frame. In the same way, double-underlined nucleotideswere removed. Lower case letters (eg. position 496 in SEQ ID 11)represent ambiguities which arose during alignment of independentsequencing reactions (some of the nucleotide sequences in the examplesare derived from combining the results of two or more experiments).

Nucleotide sequences were scanned in all six reading frames to predictthe presence of hydrophobic domains using an algorithm based on thestatistical studies of Esposti et al. [Critical evaluation of thehydropathy of membrane proteins (1990) Eur J Biochem 190:207-219]. Thesedomains represent potential transmembrane regions or hydrophobic leadersequences.

Open reading frames were predicted from fragmented nucleotide sequencesusing the program ORFFINDER (NCBI).

Underlined amino acid sequences indicate possible transmembrane domainsor leader sequences in the ORFs, as predicted by the PSORT algorithm(http://www.psort.nibb.ac.jp). Functional domains were also predictedusing the MOTIFS program (GCG Wisconsin & PROSITE).

Various tests can be used to assess the in vivo immunogencity of theproteins identified in the examples. For example, the proteins can beexpressed recombinantly and used to screen patient sera by immunoblot. Apositive reaction between the protein and patient serum indicates thatthe patient has previously mounted an immune response to the protein inquestion ie. the protein is an immunogen. This method can also be usedto identify immunodominant proteins.

The recombinant protein can also be conveniently used to prepareantibodies eg. in a mouse. These can be used for direct confirmationthat a protein is located on the cell-surface. Labelled antibody (eg.fluorescent labelling for FACS) can be incubated with intact bacteriaand the presence of label on the bacterial surface confirms the locationof the protein.

In particular, the following methods (A) to (S) were used to express,purify and biochemically characterise the proteins of the invention:

A) Chromosomal DNA Preparation

N. meningitidis strain 2996 was grown to exponential phase in 100 ml ofGC medium, harvested by centrifugation, and resuspended in 5 ml buffer(20% Sucrose, 50 mM Tris-HCl, 50 mM EDTA, pH8). After 10 minutesincubation on ice, the bacteria were lysed by adding 10 ml lysissolution (50 mM NaCl, 1% Na-Sarkosyl, 50 μg/ml Proteinase K), and thesuspension was incubated at 37° C. for 2 hours. Two phenol extractions(equilibrated to pH 8) and one ChCl₃/isoamylalcohol (24:1) extractionwere performed. DNA was precipitated by addition of 0.3M sodium acetateand 2 volumes ethanol, and was collected by centrifugation. The pelletwas washed once with 70% ethanol and redissolved in 4 ml buffer (10 mMTris-HCl, 1 mM EDTA, pH 8). The DNA concentration was measured byreading the OD at 260 nm.

B) Oligonucleotide Design

Synthetic oligonucleotide primers were designed on the basis of thecoding sequence of each ORF, using (a) the meningococcus B sequence whenavailable, or (b) the gonococcus/meningococcus A sequence, adapted tothe codon preference usage of meningococcus as necessary. Any predictedsignal peptides were omitted, by deducing the 5′-end amplificationprimer sequence immediately downstream from the predicted leadersequence.

For most ORFs, the 5′ primers included two restriction enzymerecognition sites (BamHI-NdeI, BamHI-NheI, or EcoRI-NheI, depending onthe gene's own restriction pattern); the 3′ primers included a XhoIrestriction site. This procedure was established in order to direct thecloning of each amplification product (corresponding to each ORF) intotwo different expression systems: pGEX-KG (using either BamHI-XhoI orEcoRI-XhoI), and pET21b+ (using either NdeI-XhoI or NheI-XhoI).

5′-end primer tail: CGCGGATCCCATATG (BamHI-NdeI) CGCGGATCCGCTAGC(BamHI-NheI) CCGGAATTCTAGCTAGC (EcoRI-NheI) 3′-end primer tail:CCCGCTCGAG (XhoI)

For ORFs 5, 15, 17, 19, 20, 22, 27, 28, 65 & 89, two differentamplifications were performed to clone each ORF in the two expressionsystems. Two different 5′ primers were used for each ORF; the same 3′XhoI primer was used as before:

5′-end primer tail: GGAATTCCATATGGCCATGG (NdeI) 5′-end primer tail:CGGGATCC (BamHI)

ORF 76 was cloned in the pTRC expression vector and expressed as anamino-terminus His-tag fusion. In this particular case, the predictedsignal peptide was included in the final product. NheI-BamHI restrictionsites were incorporated using primers:

5′-end primer tail: GATCAGCTAGCCATATG (NheI) 3′-end primer tail:CGGGATCC (BamHI)

As well as containing the restriction enzyme recognition sequences, theprimers included nucleotides which hybridized to the sequence to beamplified. The number of hybridizing nucleotides depended on the meltingtemperature of the whole primer, and was determined for each primerusing the formulae:

T _(m)=4(G+C)+2(A+T) (tail excluded)

T _(m)=64.9+0.41(% GC)−600/N (whole primer)

The average melting temperature of the selected oligos were 65-70° C.for the whole oligo and 50-55° C. for the hybridising region alone.

Table I (page 487) shows the forward and reverse primers used for eachamplification. In certain cases, it will be noted that the sequence ofthe primer does not exactly match the sequence in the ORF. When initialamplifications were performed, the complete 5′ and/or 3′ sequence wasnot known for some meningococcal ORFs, although the correspondingsequences had been identified in gonococcus. For amplification, thegonococcal sequences could thus be used as the basis for primer design,altered to take account of codon preference. In particular, thefollowing codons were changed: ATA→ATT; TCG→TCT; CAG→CAA; AAG→AAA;GAG→GAA; CGA→CGC; CGG→CGC; GGG→GGC. Italicised nucleotides in Table Iindicate such a change. It will be appreciated that, once the completesequence has been identified, this approach is generally no longernecessary.

Oligos were synthesized by a Perkin Elmer 394 DNA/RNA Synthesizer,eluted from the columns in 2 ml NH₄OH, and deprotected by 5 hoursincubation at 56° C. The oligos were precipitated by addition of 0.3MNa-Acetate and 2 volumes ethanol. The samples were then centrifuged andthe pellets resuspended in either 100 μl or 1 ml of water. OD₂₆₀ wasdetermined using a Perkin Elmer Lambda Bio spectophotometer and theconcentration was determined and adjusted to 2-10 pmol/μl.

C) Amplification

The standard PCR protocol was as follows: 50-200 ng of genomic DNA wereused as a template in the presence of 20-40 μM of each oligo, 400-800 μMdNTPs solution, 1×PCR buffer (including 1.5 mM MgCl₂), 2.5 units TaqIDNA polymerase (using Perkin-Elmer AmpliTaQ, GIBCO Platinum, Pwo DNApolymerase, or Tahara Shuzo Taq polymerase).

In some cases, PCR was optimsed by the addition of 10 μl DMSO or 50 μl2M betaine.

After a hot start (adding the polymerase during a preliminary 3 minuteincubation of the whole mix at 95° C.), each sample underwent adouble-step amplification: the first 5 cycles were performed using asthe hybridization temperature the one of the oligos excluding therestriction enzymes tail, followed by 30 cycles performed according tothe hybridization temperature of the whole length oligos. The cycleswere followed by a final 10 minute extension step at 72° C.

The standard cycles were as follows:

Denaturation Hybridisation Elongation First 5 cycles 30 seconds 30seconds 30-60 seconds 95° C. 50-55° C. 72° C. Last 30 cycles 30 seconds30 seconds 30-60 seconds 95° C. 65-70° C. 72° C.

The elongation time varied according to the length of the ORF to beamplified.

The amplifications were performed using either a 9600 or a 2400 PerkinElmer GeneAmp PCR System. To check the results, 1/10 of theamplification volume was loaded onto a 1-1.5% agarose gel and the sizeof each amplified fragment compared with a DNA molecular weight marker.

The amplified DNA was either loaded directly on a 1% agarose gel orfirst precipitated with ethanol and resuspended in a suitable volume tobe loaded on a 1% agarose gel. The DNA fragment corresponding to theright size band was then eluted and purified from gel, using the QiagenGel Extraction Kit, following the instructions of the manufacturer. Thefinal volume of the DNA fragment was 30 μl or 50 μl of either water or10 mM Tris, pH 8.5.

D) Digestion of PCR Fragments

The purified DNA corresponding to the amplified fragment was split into2 aliquots and double-digested with:

-   -   NdeI/XhoI or NheI/XhoI for cloning into pET-21b+ and further        expression of the protein as a C-terminus His-tag fusion    -   BamHI/XhoI or EcoRI/XhoI for cloning into pGEX-KG and further        expression of the protein as N-terminus GST fusion.    -   For ORF 76, NheI/BamHI for cloning into pTRC-H isA vector and        further expression of the protein as N-terminus His-tag fusion.    -   EcoRI/PstI, EcoRI/SalI, SalI/PstI for cloning into pGex-His and        further expression of the protein as N-terminus His-tag fusion

Each purified DNA fragment was incubated (37° C. for 3 hours toovernight) with 20 units of each restriction enzyme (New EnglandBiolabs) in a either 30 or 40 μl final volume in the presence of theappropriate buffer. The digestion product was then purified using theQIAquick PCR purification kit, following the manufacturer'sinstructions, and eluted in a final volume of 30 or 50 μl of eitherwater or 10 mM Tris-HCl, pH 8.5. The final DNA concentration wasdetermined by 1% agarose gel electrophoresis in the presence of titratedmolecular weight marker.

E) Digestion of the Cloning Vectors (pET22B, pGEX-KG, pTRC-His A, andpGex-His)

10 μg plasmid was double-digested with 50 units of each restrictionenzyme in 200 μl reaction volume in the presence of appropriate bufferby overnight incubation at 37° C. After loading the whole digestion on a1% agarose gel, the band corresponding to the digested vector waspurified from the gel using the Qiagen QIAquick Gel Extraction Kit andthe DNA was eluted in 50 μl of 10 mM Tris-HCl, pH 8.5. The DNAconcentration was evaluated by measuring OD₂₆₀ of the sample, andadjusted to 50 μg/μl. 1 μl of plasmid was used for each cloningprocedure.

The vector pGEX-His is a modified pGEX-2T vector carrying a regionencoding six histidine residues upstream to the thrombin cleavage siteand containing the multiple cloning site of the vector pTRC99(Pharmacia).

F) Cloning

The fragments corresponding to each ORF, previously digested andpurified, were ligated in both pET22b and pGEX-KG. In a final volume of20 μl, a molar ratio of 3:1 fragment/vector was ligated using 0.5 μl ofNEB T4 DNA ligase (400 units/μl), in the presence of the buffer suppliedby the manufacturer. The reaction was incubated at room temperature for3 hours. In some experiments, ligation was performed using theBoheringer “Rapid Ligation Kit”, following the manufacturer'sinstructions.

In order to introduce the recombinant plasmid in a suitable strain, 100μl E. coli DH5 competent cells were incubated with the ligase reactionsolution for 40 minutes on ice, then at 37° C. for 3 minutes, then,after adding 800 μl LB broth, again at 37° C. for 20 minutes. The cellswere then centrifuged at maximum speed in an Eppendorf microfuge andresuspended in approximately 200 μl of the supernatant. The suspensionwas then plated on LB ampicillin (100 mg/ml).

The screening of the recombinant clones was performed by growing 5randomly-chosen colonies overnight at 37° C. in either 2 ml (pGEX or pTCclones) or 5 ml (pET clones) LB broth+100 μg/ml ampicillin. The cellswere then pelletted and the DNA extracted using the Qiagen QIAprep SpinMiniprep Kit, following the manufacturer's instructions, to a finalvolume of 30 μl. 5 μl of each individual miniprep (approximately 1 g)were digested with either NdeI/XhoI or BamHI/XhoI and the wholedigestion loaded onto a 1-1.5% agarose gel (depending on the expectedinsert size), in parallel with the molecular weight marker (1 Kb DNALadder, GIBCO). The screening of the positive clones was made on thebase of the correct insert size.

For the cloning of ORFs 110, 111, 113, 115, 119, 122, 125 & 130, thedouble-digested PCR product was ligated into double-digested vectorusing EcoRI-PstI cloning sites or, for ORFs 115 & 127, EcoRI-SalI or,for ORF 122, SalI-PstI. After cloning, the recombinant plasmids wereintroduced in the E. coli host W3110. Individual clones were grownovernight at 37° C. in L-broth with 50 μl/ml ampicillin.

G) Expression

Each ORF cloned into the expression vector was transformed into thestrain suitable for expression of the recombinant protein product. 1 μlof each construct was used to transform 30 μl of E. coli BL21 (pGEXvector), E. coli TOP 10 (pTRC vector) or E. coli BL21-DE3 (pET vector),as described above. In the case of the pGEX-His vector, the same E. colistrain (W3110) was used for initial cloning and expression. Singlerecombinant colonies were inoculated into 2 ml LB+Amp (100 μg/ml),incubated at 37° C. overnight, then diluted 1:30 in 20 ml of LB+Amp (100μg/ml) in 100 ml flasks, making sure that the OD₆₀₀ ranged between 0.1and 0.15. The flasks were incubated at 30° C. into gyratory water bathshakers until OD indicated exponential growth suitable for induction ofexpression (0.4-0.8 OD for pET and pTRC vectors; 0.8-1 OD for pGEX andpGEX-His vectors). For the pET, pTRC and pGEX-His vectors, the proteinexpression was induced by addition of 1 mM IPTG, whereas in the case ofpGEX system the final concentration of IPTG was 0.2 mM. After 3 hoursincubation at 30° C., the final concentration of the sample was checkedby OD. In order to check expression, 1 ml of each sample was removed,centrifuged in a microfuge, the pellet resuspended in PBS, and analysedby 12% SDS-PAGE with Coomassie Blue staining. The whole sample wascentrifuged at 6000 g and the pellet resuspended in PBS for further use.

H) GST-Fusion Proteins Large-Scale Purification.

A single colony was grown overnight at 37° C. on LB+Amp agar plate. Thebacteria were inoculated into 20 ml of LB+Amp liquid colture in a waterbath shaker and grown overnight. Bacteria were diluted 1:30 into 600 mlof fresh medium and allowed to grow at the optimal temperature (20-37°C.) to OD₅₅₀ 0.8-1. Protein expression was induced with 0.2 mM IPTGfollowed by three hours incubation. The culture was centrifuged at 8000rpm at 4° C. The supernatant was discarded and the bacterial pellet wasresuspended in 7.5 ml cold PBS. The cells were disrupted by sonicationon ice for 30 sec at 40 W using a Branson sonifier B-15, frozen andthawed twice and centrifuged again. The supernatant was collected andmixed with 150 μl Glutatione-Sepharose 4B resin (Pharmacia) (previouslywashed with PBS) and incubated at room temperature for 30 minutes. Thesample was centrifuged at 700 g for 5 minutes at 4° C. The resin waswashed twice with 10 ml cold PBS for 10 minutes, resuspended in 1 mlcold PBS, and loaded on a disposable column. The resin was washed twicewith 2 ml cold PBS until the flow-through reached OD₂₈₀ of 0.02-0.06.The GST-fusion protein was eluted by addition of 700 μl cold Glutathioneelution buffer (10 mM reduced glutathione, 50 mM Tris-HCl) and fractionscollected until the OD₂₈₀ was 0.1.21 μl of each fraction were loaded ona 12% SDS gel using either Biorad SDS-PAGE Molecular weight standardbroad range (M1) (200, 116.25, 97.4, 66.2, 45, 31, 21.5, 14.4, 6.5 kDa)or Amersham Rainbow Marker (M2) (220, 66, 46, 30, 21.5, 14.3 kDa) asstandards. As the MW of GST is 26 kDa, this value must be added to theMW of each GST-fusion protein.

I) His-Fusion Solubility Analysis (ORFs 111-129)

To analyse the solubility of the His-fusion expression products, pelletsof 3 ml cultures were resuspended in buffer M1 [500 μl PBS pH 7.2]. 25μl lysozyme (10 mg/ml) was added and the bacteria were incubated for 15min at 4° C. The pellets were sonicated for 30 sec at 40 W using aBranson sonifier B-15, frozen and thawed twice and then separated againinto pellet and supernatant by a centrifugation step. The supernatantwas collected and the pellet was resuspended in buffer M2 [8M urea, 0.5MNaCl, 20 mM imidazole and 0.1 M NaH₂ PO₄] and incubated for 3 to 4 hoursat 4° C. After centrifugation, the supernatant was collected and thepellet was resuspended in buffer M3 [6M guanidinium-HCl, 0.5M NaCl, 20mM imidazole and 0.1M NaH₂PO₄] overnight at 4° C. The supernatants fromall steps were analysed by SDS-PAGE.

The proteins expressed from ORFs 113, 119 and 120 were found to besoluble in PBS, whereas ORFs 111, 122, 126 and 129 need urea and ORFs125 and 127 need guanidium-HCl for their solubilization.

J) His-Fusion Large-Scale Purification.

A single colony was grown overnight at 37° C. on a LB+Amp agar plate.The bacteria were inoculated into 20 ml of LB+Amp liquid culture andincubated overnight in a water bath shaker. Bacteria were diluted 1:30into 600 ml fresh medium and allowed to grow at the optimal temperature(20-37° C.) to OD₅₅₀ 0.6-0.8. Protein expression was induced by additionof 1 mM IPTG and the culture further incubated for three hours. Theculture was centrifuged at 8000 rpm at 4° C., the supernatant wasdiscarded and the bacterial pellet was resuspended in 7.5 ml of either(i) cold buffer A (300 mM NaCl, 50 mM phosphate buffer, 10 mM imidazole,pH 8) for soluble proteins or (ii) buffer B (urea 8M, 10 mM Tris-HCl,100 mM phosphate buffer, pH 8.8) for insoluble proteins. The cells weredisrupted by sonication on ice for 30 sec at 40 W using a Bransonsonifier B-15, frozen and thawed two times and centrifuged again.

For insoluble proteins, the supernatant was stored at −20° C., while thepellets were resuspended in 2 ml buffer C (6M guanidine hydrochloride,100 mM phosphate buffer, 10 mM Tris-HCl, pH 7.5) and treated in ahomogenizer for 10 cycles. The product was centrifuged at 13000 rpm for40 minutes.

Supernatants were collected and mixed with 150 μl Ni²⁺-resin (Pharmacia)(previously washed with either buffer A or buffer B, as appropriate) andincubated at room temperature with gentle agitation for 30 minutes. Thesample was centrifuged at 700 g for 5 minutes at 4° C. The resin waswashed twice with 10 ml buffer A or B for 10 minutes, resuspended in 1ml buffer A or B and loaded on a disposable column. The resin was washedat either (i) 4° C. with 2 ml cold buffer A or (ii) room temperaturewith 2 ml buffer B, until the flow-through reached OD₂₈₀ of 0.02-0.06.

The resin was washed with either (i) 2 ml cold 20 mM imidazole buffer(300 mM NaCl, 50 mM phosphate buffer, 20 mM imidazole, pH 8) or (ii)buffer D (urea 8M, 10 mM Tris-HCl, 100 mM phosphate buffer, pH 6.3)until the flow-through reached the OD₂₈₀ of 0.02-0.06. The His-fusionprotein was eluted by addition of 700 μl of either (i) cold elutionbuffer A (300 mM NaCl, 50 mM phosphate buffer, 250 mM imidazole, pH 8)or (ii) elution buffer B (urea 8M, 10 mM Tris-HCl, 100 mM phosphatebuffer, pH 4.5) and fractions collected until the OD₂₈₀ was 0.1. 21 μlof each fraction were loaded on a 12% SDS gel.

K) His-Fusion Proteins Renaturation

10% glycerol was added to the denatured proteins. The proteins were thendiluted to 20 μg/ml using dialysis buffer I (10% glycerol, 0.5Marginine, 50 mM phosphate buffer, 5 mM reduced glutathione, 0.5 mMoxidised glutathione, 2M urea, pH 8.8) and dialysed against the samebuffer at 4° C. for 12-14 hours. The protein was further dialysedagainst dialysis buffer II (10% glycerol, 0.5M arginine, 50 mM phosphatebuffer, 5 mM reduced glutathione, 0.5 mM oxidised glutathione, pH 8.8)for 12-14 hours at 4° C. Protein concentration was evaluated using theformula:

Protein (mg/ml)=(1.55×OD₂₈₀)−(0.76×OD₂₆₀)

L) His-Fusion Large-Scale Purification (ORFs 111-129)

500 ml of bacterial cultures were induced and the fusion proteins wereobtained soluble in buffer M1, M2 or M3 using the procedure describedabove. The crude extract of the bacteria was loaded onto a Ni-NTAsuperflow column (Quiagen) equilibrated with buffer M1, M2 or M3depending on the solubilization buffer of the fusion proteins. Unboundmaterial was eluted by washing the column with the same buffer. Thespecific protein was eluted with the corresponding buffer containing 500mM imidazole and dialysed against the corresponding buffer withoutimidazole. After each run the columns were sanitized by washing with atleast two column volumes of 0.5 M sodium hydroxide and reequilibratedbefore the next use.

M) Mice Immunisations

20 μg of each purified protein were used to immunise miceintraperitoneally. In the case of ORFs 2, 4, 15, 22, 27, 28, 37, 76, 89and 97, Balb-C mice were immunised with Al(OH)₃ as adjuvant on days 1,21 and 42, and immune response was monitored in samples taken on day 56.For ORFs 44, 106 and 132, CD1 mice were immunised using the sameprotocol. For ORFs 25 and 40, CD1 mice were immunised using Freund'sadjuvant, rather than AL(OH)₃, and the same immunisation protocol wasused, except that the immune response was measured on day 42, ratherthan 56. Similarly, for ORFs 23, 32, 38 and 79, CD1 mice were immunisedwith Freund's adjuvant, but the immune response was measured on day 49.

N) ELISA Assay (Sera Analysis)

The acapsulated MenB M7 strain was plated on chocolate agar plates andincubated overnight at 37° C. Bacterial colonies were collected from theagar plates using a sterile dracon swab and inoculated into 7 ml ofMueller-Hinton Broth (Difco) containing 0.25% Glucose. Bacterial growthwas monitored every 30 minutes by following OD₆₂₀. The bacteria were letto grow until the OD reached the value of 0.3-0.4. The culture wascentrifuged for 10 minutes at 10000 rpm. The supernatant was discardedand bacteria were washed once with PBS, resuspended in PBS containing0.025% formaldehyde, and incubated for 2 hours at room temperature andthen overnight at 4° C. with stirring. 100 μl bacterial cells were addedto each well of a 96 well Greiner plate and incubated overnight at 4° C.The wells were then washed three times with PBT washing buffer (0.1%Tween-20 in PBS). 200 μl of saturation buffer (2.7% Polyvinylpyrrolidone10 in water) was added to each well and the plates incubated for 2 hoursat 37° C. Wells were washed three times with PBT. 200 μl of diluted sera(Dilution buffer: 1% BSA, 0.1% Tween-20, 0.1% NaN₃ in PBS) were added toeach well and the plates incubated for 90 minutes at 37° C. Wells werewashed three times with PBT. 100 μl of HRP-conjugated rabbit anti-mouse(Dako) serum diluted 1:2000 in dilution buffer were added to each welland the plates were incubated for 90 minutes at 37° C. Wells were washedthree times with PBT buffer. 100 μl of substrate buffer for HRP (25 mlof citrate buffer pH5, 10 mg of O-phenildiamine and 10 μl of H₂O) wereadded to each well and the plates were left at room temperature for 20minutes. 100 μl H₂SO₄ was added to each well and OD₄₉₀ was followed. TheELISA was considered positive when OD₄₉₀ was 2.5 times the respectivepre-immune sera.

O) FACScan Bacteria Binding Assay Procedure.

The acapsulated MenB M7 strain was plated on chocolate agar plates andincubated overnight at 37° C. Bacterial colonies were collected from theagar plates using a sterile dracon swab and inoculated into 4 tubescontaining 8 ml each Mueller-Hinton Broth (Difco) containing 0.25%glucose. Bacterial growth was monitored every 30 minutes by followingOD₆₂₀. The bacteria were let to grow until the OD reached the value of0.35-0.5. The culture was centrifuged for 10 minutes at 4000 rpm. Thesupernatant was discarded and the pellet was resuspended in blockingbuffer (1% BSA, 0.4% NaN₃) and centrifuged for 5 minutes at 4000 rpm.Cells were resuspended in blocking buffer to reach OD₆₂₀ of 0.07. 100 μlbacterial cells were added to each well of a Costar 96 well plate. 100μl of diluted (1:200) sera (in blocking buffer) were added to each welland plates incubated for 2 hours at 4° C. Cells were centrifuged for 5minutes at 4000 rpm, the supernatant aspirated and cells washed byaddition of 200 μl/well of blocking buffer in each well. 100 μl ofR-Phicoerytrin conjugated F(ab)₂ goat anti-mouse, diluted 1:100, wasadded to each well and plates incubated for 1 hour at 4° C. Cells werespun down by centrifugation at 4000 rpm for 5 minutes and washed byaddition of 200 μl/well of blocking buffer. The supernatant wasaspirated and cells resuspended in 200 μl/well of PBS, 0.25%formaldehyde. Samples were transferred to FACScan tubes and read. Thecondition for FACScan setting were: FL1 on, FL2 and FL3 off; FSC-Hthreshold:92; FSC PMT Voltage: E 02; SSC PMT: 474; Amp. Gains 7.1; FL-2PMT: 539; compensation values: 0.

P) OMV Preparations

Bacteria were grown overnight on 5 GC plates, harvested with a loop andresuspended in 10 ml 20 mM Tris-HCl. Heat inactivation was performed at56° C. for 30 minutes and the bacteria disrupted by sonication for 10minutes on ice (50% duty cycle, 50% output). Unbroken cells were removedby centrifugation at 5000 g for 10 minutes and the total cell envelopefraction recovered by centrifugation at 50000 g at 4° C. for 75 minutes.To extract cytoplasmic membrane proteins from the crude outer membranes,the whole fraction was resuspended in 2% sarkosyl (Sigma) and incubatedat room temperature for 20 minutes. The suspension was centrifuged at10000 g for 10 minutes to remove aggregates, and the supernatant furtherultracentrifuged at 50000 g for 75 minutes to pellet the outermembranes. The outer membranes were resuspended in 10 mM Tris-HCl, pH8and the protein concentration measured by the Bio-Rad Protein assay,using BSA as a standard.

Q) Whole Extracts Preparation

Bacteria were grown overnight on a GC plate, harvested with a loop andresuspended in 1 ml of 20 mM Tris-HCl. Heat inactivation was performedat 56° C. for 30 minutes.

R) Western Blotting

Purified proteins (500 ng/lane), outer membrane vesicles (5 μg) andtotal cell extracts (25 μg) derived from MenB strain 2996 were loaded on15% SDS-PAGE and transferred to a nitrocellulose membrane. The transferwas performed for 2 hours at 150 mA at 4° C., in transferring buffer(0.3% Tris base, 1.44% glycine, 20% methanol). The membrane wassaturated by overnight incubation at 4° C. in saturation buffer (10%skimmed milk, 0.1% Triton X100 in PBS). The membrane was washed twicewith washing buffer (3% skimmed milk, 0.1% Triton X100 in PBS) andincubated for 2 hours at 37° C. with mice sera diluted 1:200 in washingbuffer. The membrane was washed twice and incubated for 90 minutes witha 1:2000 dilution of horseradish peroxidase labelled anti-mouse Ig. Themembrane was washed twice with 0.1% Triton X100 in PBS and developedwith the Opti-4CN Substrate Kit (Bio-Rad). The reaction was stopped byadding water.

S) Bactericidal Assay

MC58 strain was grown overnight at 37° C. on chocolate agar plates. 5-7colonies were collected and used to inoculate 7 ml Mueller-Hinton broth.The suspension was incubated at 37° C. on a nutator and let to growuntil OD₆₂₀ was 0.5-0.8. The culture was aliquoted into sterile 1.5 mlEppendorf tubes and centrifuged for 20 minutes at maximum speed in amicrofuge. The pellet was washed once in Gey's buffer (Gibco) andresuspended in the same buffer to an OD₆₂₀ of 0.5, diluted 1:20000 inGey's buffer and stored at 25° C.

50 μl of Gey's buffer/]% BSA was added to each well of a 96-well tissueculture plate. 25 μl of diluted mice sera (1:100 in Gey's buffer/0.2%BSA) were added to each well and the plate incubated at 4° C. 25 μl ofthe previously described bacterial suspension were added to each well.25 μl of either heat-inactivated (56° C. waterbath for 30 minutes) ornormal baby rabbit complement were added to each well. immediately afterthe addition of the baby rabbit complement, 22 μl of each sample/wellwere plated on Mueller-Hinton agar plates (time 0). The 96-well platewas incubated for 1 hour at 37° C. with rotation and then 22 μl of eachsample/well were plated on Mueller-Hinton agar plates (time 1). Afterovernight incubation the colonies corresponding to time 0 and time 1hour were counted.

Table II (page 493) gives a summary of the cloning, expression andprurification results.

Example 1

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 1>:

1 ATGAAACAGA CAGTCAA.AT GCTTGCCGCC GCCCTGATTG CCTTGGGCTT 51 GAACCGACCGGTGTGGNCGG ATGACGTATC GGATTTTCGG GAAAACTTGC 101 A.GCGGCAGC ACAGGGAAATGCAGCAGCCC AATACAATTT GGGCGCAATG 151 TAT.TACAAA GGACGCGCGT GCGCCGGGATGATGCTGAAG CGGTCAGATG 201 GTATCGGCAG CCGGCGGAAC AGGGGTTAGC CCAAGCCCAATACAATTTGG 251 GCTGGATGTA TGCCAACGGG CGCGC.GTGC GCCAAGATGA TACCGAAGCG301 GTCAGATGGT ATCGGCAGGC GGCAGCGCAG GGGGTTGTCC AAGCCCAATA 351CAATTTGGGC GTGATATATG CCGAAGGACG TGGAGTGCGC CAAGACGATG 401 TCGAAGCGGTCAGATGGTTT CGGCAGGCGG CAGCGCAGGG GGTAGCCCAA 451 GCCCAAAACA ATTTGGGCGTGATGTATGCC GAAAGANCGC GCGTGCGCCA 501 AGACCG...

This corresponds to the amino acid sequence <SEQ ID 2; ORF37>:

1 MKQTVXMLAA ALIALGLNRP VWXDDVSDFR ENLXAAAQGN AAAQYNLGAM 51 YXQRTRVRRDDAEAVRWYRQ PAEQGLAQAQ YNLGWMYANG RXVRQDDTEA 101 VRWYRQAAAQ GVVQAQYNLGVIYAEGRGVR QDDVEAVRWF RQAAAQGVAQ 151 AQNNLGVMYA ERXRVRQD...

Further work revealed the complete nucleotide sequence <SEQ ID 3>:

1 ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG CCTTGGGCTT 51 GAACCGAGCGGTGTGGGCGG ATGACGTATC GGATTTTCGG GAAAACTTGC 101 AGGCGGCAGC ACAGGGAAATGCAGCAGCCC AATACAATTT GGGCGCAATG 151 TATTACAAAG GACGCGGCGT GCGCCGGGATGATGCTGAAG CGGTCAGATG 201 GTATCGGCAG GCGGCGGAAC AGGGGTTAGC CCAAGCCCAATACAATTTGG 251 GCTGGATGTA TGCCAACGGG CGCGGCGTGC GCCAAGATGA TACCGAAGCG301 GTCAGATGGT ATCGGCAGGC GGCAGCGCAG GGGGTTGTCC AAGCCCAATA 351CAATTTGGGC GTGATATATG CCGAAGGACG TGGAGTGCGC CAAGACGATG 401 TCGAAGCGGTCAGATGGTTT CGGCAGGCGG CAGCGCAGGG GGTAGCCCAA 451 GCCCAAAACA ATTTGGGCGTGATGTATGCC GAAAGACGCG GCGTGCGCCA 501 AGACCGCGCC CTTGCACAAG AATGGTTTGGCAAGGCTTGT CAAAACGGAG 551 ACCAAGACGG CTGCGACAAT GACCAACGCC TGAAGGCGGGTTATTGA

This corresponds to the amino acid sequence <SEQ ID 4; ORF37-1>:

1 MKQTVKWLAA ALIALGLNRA VWADDVSDFR ENLQAAAQGN AAAQYNLGAM 51 YYKGRGVRRDDAEAVRWYRQ AAEQGLAQAQ YNLGWMYANG RGVRQDDTEA 101 VRWYRQAAAQ GVVQAQYNLGVIYAEGRGVR QDDVEAVRWF RQAAAQGVAQ 151 AQNNLGVMYA ERRGVRQDRA LAQEWFGKACQNGDQDGCDN DQRLKAGY*

Further work identified the corresponding gene in strain A of N.meningitidis <SEQ ID 5>:

1 ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG CCTTGGGCTT 51 GAACCAAGCGGTGTGGGCGG ATGACGTATC GGATTTTCGG GAAAACTTGC 101 AGGCGGCAGC ACAGGGAAATGCAGCAGCCC AAAACAATTT GGGCGTGATG 151 TATGCCGAAA GACGCGGCGT GCGCCAAGACCGCGCCCTTG CACAAGAATG 201 GCTTGGCAAG GCTTGTCAAA ACGGATACCA AGACAGCTGCGACAATGACC 251 AACGCCTGAA AGCGGGTTAT TGA

This encodes a protein having amino acid sequence <SEQ ID 6; ORF37a>:

1 MKQTVKWLAA ALIALGLNQA VWADDVSDFR ENLQAAAQGN AAAQNNLGVM 51 YAERRGVRQDRALAQEWLGK ACQNGYQDSC DNDQRLKAGY *

The originally-identified partial strain B sequence (ORF37) shows 68.0%identity over a 75aa overlap with ORF37a:

Further work identified the corresponding gene in N. gonorrhoeae <SEQ ID7>:

1 ATGAAACAGA CAGTCAAATG GCTTGCCGCC GCCCTGATTG CCTTGGGCTT 51 GAACCAAGCGGTGTGGGCGG GTGACGTATC GGATTTTCGG GAAAACTTGC 101 AGgcggcaGA ACaggGAAATGCAGCAGCCC AATTCAATTT GGGCGTGATG 151 TATGAAAATG GACAAGGAGT TCGTCAAGATTATGTACAGG CAGTGCAGTG 201 GTATCGCAAG GCTTCAGAAC AAGGGGATGC CCAAGCCCAATACAATTTGG 251 GCTTGATGTA TTACGATGGA CGCGGCGTGC GCCAAGACCT TGCGCTCGCT301 CAACAATGGC TTGGCAAGGC TTGTCAAAAC GGAGACCAAA ACAGCTGCGA 351CAATGACCAA CGCCTGAAGG CGGGTTATTA A

This encodes a protein having amino acid sequence <SEQ ID 8; ORF37ng>:

1 MKQTVKWLAA ALIALGLNQA VWAGDVSDFR ENLQAAEQGN AAAQFNLGVM 51 YENGQGVRQDYVQAVQWYRK ASEQGDAQAQ YNLGLMYYDG RGVRQDLALA 101 QQWLGKACQN GDQNSCDNDQRLKAGY*

The originally-identified partial strain B sequence (ORF37) shows 64.9%identity over a 111aa overlap with ORF37ng:

The complete strain B sequence (ORF37-1) and ORF37ng show 51.5% identityin 198 aa overlap:

Computer analysis of these amino acid sequences indicates a putativeleader sequence, and it was predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF37-1 (11 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 1A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 1B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for ELISA(positive result), FACS analysis (FIG. 1C), and a bactericidal assay(FIG. 1D). These experiments confirm that ORF37-1 is a surface-exposedprotein, and that it is a useful immunogen.

FIG. 1E shows plots of hydrophilicity, antigenic index, and AMPHIregions for ORF37-1.

Example 2

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 9>:

  TTCGGCGA CATCGGCGGT TTGAAGGTCA ATGCCCCCGTCAAATCCGCA GGCGTATTGG TCGGGCGCGT CGGCGCTATCGGACTTGACC CGAAATCCTA TCAGGCGAGG GTGCGCCTCGATTTGGACGG CAAGTATCAG TTCAGCAGCG ACGTTTCCGCGCAAATCCTG ACTTCsGGAC TTTTGGGCGA GCAGTACATCGGGCTGCAGC AGGGCGGCGA CACGGAAAAC CTTGCTGCCGGCGACACCAT CTCCGTAACC AGTTCTGCAA TGGTTCTGGAAAACCTTATC GGCAAATTCA TGACGAGTTT TGCCGAGAAAAATGCCGACG GCGGCAATGC GGAAAAAGCC GCCGAATAA

This corresponds to the amino acid sequence <SEQ ID 10>:

  1 FGDIGGLKVN APVKSAGVLV GRVGAIGLDP KSYQARVRLD LDGKYQFSSD 51 VSAQILTSGL LGEQYIGLQQ GGDTENLAAG DTISVTSSAM VLENLIGKFM101 TSFAEKNADG GNAEKAAE*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Hypothetical H. influenzae Protein (ybrd.haein;Accession Number p45029)

SEQ ID 9 and ybrd.haein show 48.4% aa identity in 122 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

SEQ ID 9 shows 99.2% identity over a 118aa overlap with a predicted ORFfrom N. gonorrhoeae:

The complete yrbd H. influenzae sequence has a leader sequence and it isexpected that the full-length homologous N. meningitidis protein willalso have one. This suggests that it is either a membrane protein, asecreted protein, or a surface protein and that the protein, or one ofits epitopes, could be a useful antigen for vaccines or diagnostics.

Example 3

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 11>:

1 ..ATTTTGATAT ACCTCATCCG CAAGAATCTA GGTTCGCCCG TCTTCTTCTT 51  TCAGGAACGC CCCGGAAAGG ACGGAAAACC TTTTAAAATG GTCAAATTCC 101  GTTCCATGCG CGACGGCTTG TATTCAGACG GCATTCCGCT GCCCGACGGA 151  GAACGCCTGA CACCGTTCGG CAAAAAACTG CGTGCCGcCA GTwTGGACGA 201  ACTGCCTGAA TTATGGAATA TCTTAAAAGG CGAGATGAGC CTGGTCGGCC 251  CCCGCCCGCT GCTGATGCAA TATCTGCCGC TGTACGACAA CTTCCAAAAC 301  CGCCGCCACG AAATGAAACC CGGCATTACC GGCTGGGCGC AGGTCAACGG 351  GCGCAACGCg CTTTCGTGGG ACGAAAAATT CGCCTGCGAT GTTTGGTATA 401  TCGACCACTT CAGCCTGTGC CTCGACATCA AAATCCTACT GCTGACGGTT 451  AAAAAAGTAT TAATCAAGGA AGGGATTTCC GCACAGGGCG AACA.aCCAT 501  GCCCCCTTTC ACAGGAAAAC GCAAACTCGC CGTCGTCGGT GCGGGCGGAC 551  ACGGAAAAGT CGTTGCCGAC CTTGCCGCCG CACTCGGCCG GTACAGGGAA 601  ATCGTTTTTC TGGACGACCG CGCACAAGGC AGCGTCAACG GCTTTTCCGT 651  CATCGGCACG ACGCTGCTGC TTGAAAACAG TTTATCGCCC GAACAATACG 701  ACGTCGCCGT CGCCGTCGGC AACAACCGCA TCCGCCGCCA AATCGCCGAA 751  AAAGCCGCCG CGCTCGGCTT CGCCCTGCCC GTACTGGTTC ATCCGGACGC 801  GACCGTCTCG CCTTCTGCAA CAGTCGGACA AGGCAGCGTC GTTATGGCGA 851  AAGCGGTCG..

This corresponds to the amino acid sequence <SEQ ID 12; ORF3>:

1 . . . ILIYLIRKNL GSPVFFFQER PGKDGKPFKM VKFRSMRDGL YSDGIPLPDG 51      ERLTPFGKKL RAASXDELPE LWNILKGEMS LVGPRPLLMQ YLPLYDNFQN 101      RRHEMKPGIT GWAQVNGRNA LSWDEKFACD VWYIDHFSLC LDIKILLLTV 151      KKVLIKEGIS AQGEXTMPPF TGKRKLAVVG AGGHGKVVAD LAAALGRYRE 201      IVFLDDRAQG SVNGFSVIGT TLLLENSLSP EQYDVAVAVG NNRIRRQIAE 251      KAAALGFALP VLVHPDATVS PSATVGQGSV VMAKAV . . .

Further sequence analysis revealed the complete nucleotide sequence <SEQID 13>:

1 ATGAGTAAAT TCTTCAAACG CCTGTTTGAC ATTGTTGCCT CCGCCTCGGG 51ACTGATTTTC CTCTCGCCAG TATTTTTGAT TTTGATATAC CTCATCCGCA 101AGAATCTAGG TTCGCCCGTC TTCTTCTTTC AGGAACGCCC CGGAAAGGAC 151GGAAAACCTT TTAAAATGGT CAAATTCCGT TCCATGCGCG ACGCGCTTGA 201TTCAGACGGC ATTCCGCTGC CCGACGGAGA ACGCCTGACA CCGTTCGGCA 251AAAAACTGCG TGCCGCCAGT TTGGACGAAC TGCCTGAATT ATGGAATATC 301TTAAAAGGCG AGATGAGCCT GGTCGGCCCC CGCCCGCTGC TGATGCAATA 351TCTGCCGCTG TACGACAACT TCCAAAACCG CCGCCACGAA ATGAAACCCG 401GCATTACCGG CTGGGCGCAG GTCAACGGGC GCAACGCGCT TTCGTGGGAC 451GAAAAATTCG CCTGCGATGT TTGGTATATC GACCACTTCA GCCTGTGCCT 501CGACATCAAA ATCCTACTGC TGACGGTTAA AAAAGTATTA ATCAAGGAAG 551GGATTTCCGC ACAGGGCGAA GCCACCATGC CCCCTTTCAC AGGAAAACGC 601AAACTCGCCG TCGTCGGTGC GGGCGGACAC GGAAAAGTCG TTGCCGACCT 651TGCCGCCGCA CTCGGCCGGT ACAGGGAAAT CGTTTTTCTG GACGACCGCG 701CACAAGGCAG CGTCAACGGC TTTTCCGTCA TCGGCACGAC GCTGCTGCTT 751GAAAACAGTT TATCGCCCGA ACAATACGAC GTCGCCGTCG CCGTCGGCAA 801CAACCGCATC CGCCGCCAAA TCGCCGAAAA AGCCGCCGCG CTCGGCTTCG 851CCCTGCCCGT TCTGGTTCAT CCGGACGCGA CCGTCTCGCC TTCTGCAACA 901GTCGGACAAG GCAGCGTCGT TATGGCGAAA GCCGTCGTAC AGGCAGGCAG 951CGTATTGAAA GACGGCGTGA TTGTGAACAC TGCCGCCACC GTCGATCACG 1001ACTGCCTGCT TAACGCTTTC GTCCACATCA GCCCAGGCGC GCACCTGTCG 1051GGCAACACGC ATATCGGCGA AGAAAGCTGG ATAGGCACGG GCGCGTGCAG 1101CCGCCAGCAG ATCCGTATCG GCAGCCGCGC AACCATTGGA GCGGGCGCAG 1151TCGTCGTACG CGACGTTTCA GACGGCATGA CCGTCGCGGG CAATCCGGCA 1201AAGCCGCTGC CGCGCAAAAA CCCCGAGACC TCGACAGCAT AA

This corresponds to the amino acid sequence <SEQ ID 14; ORF3-1>:

1 MSKFFKRLFD IVASASGLIF LSPVFLILIY LIRKNLGSPV FFFQERPGKD 51GKPFKMVKFR SMRDALDSDG IPLPDGERLT PFGKKLRAAS LDELPELWNI 101LKGEMSLVGP RPLLMQYLPL YDNFQNRRHE MKPGITGWAQ VNGRNALSWD 151EKFACDVWYI DHFSLCLDIK ILLLTVKKVL IKEGISAQGE ATMPPFTGKR 201KLAVVGAGGH GKVVADLAAA LGRYREIVFL DDRAQGSVNG FSVIGTTLLL 251ENSLSPEQYD VAVAVGNNRI RRQIAEKAAA LGFALPVLVH PDATVSPSAT 301VGQGSVVMAK AVVQAGSVLK DGVIVNTAAT VDHDCLLNAF VHISPGAHLS 351GNTHIGEESW IGTGACSRQQ IRIGSRATIG AGAVVVRDVS DGMTVAGNPA 401KPLPRKNPET STA*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF3 shows 93.0% identity over a 286aa overlap with an ORF (ORF3a) fromstrain A of N. meningitidis:

The complete length ORF3a nucleotide sequence <SEQ ID 15> is:

1 ATGAGTAAAT TCTTCAAACG CCTGTTTGAC ATTGTTGCCT CCGCCTCGGG 51 ACTGATTTTCCTCTCGCCAG TATTTTTGAT TTTGATATAC CTCATCCGCA 101 AGAATCTGGG TTCGCCCGTCTTCTTCTTTC AGGAACGCCC CGGAAAGGAC 151 GGAAAACCTT TTAAAATGGT CAAATTCCGTTCCATGCACG ACGCGCTTGA 201 TTCAGACGGC ATTCTGCTGC CCGACGGAGA ACGCCTGACACCGTTCGGCA 251 AAAAACTGCG TGCCGCCAGT TTGGACGAAC TGCCCGAACT GTGGAACGTC301 CTCAAAGGCG ACATGAGCCT GGTCGGCCCC CGCCCGCTGC TGATGCAATA 351TCTGCCGCTG TACGACAACT TCCAAAACCG CCGCCACGAA ATGAAACCGG 401 GCATTACCGGCTGGGCGCAG GTCAACGGGC GCAACGCGCT TTCGTGGGAC 451 GAACGCTTCG CATGCGACATCTGGTATATC GACCACTTCA GCCTGTGCCT 501 CGAGATCAAA ATCCTACTGC TGACGGTTAAAAAAGTATTA ATCAAAGAAG 551 GGATTTCCGC ACAGGGCGAA GCCACCATGC CCCCTTTCACAGGAAAACGC 601 AAACTTGCCG TCGTCGGTGC GGGCGGACAC GGCAAAGTCG TTGCCGAGCT651 TGCCGCCGCA CTCGGCACAT ACGGCGAAAT CGTTTTTCTG GACGACCGCG 701TCCAAGGCAG CGTCAACGGC TTCCCCGTCA TCGGCACGAC GCTGCTGCTT 751 GAAAACAGTTTATCGCCCGA ACAATTCGAC ATCGCCGTCG CCGTCGGCAA 801 CAACCGCATC CGCCGCCAAATCGCCGAAAA AGCCGCCGCG CTCGGCTTCG 851 CCCTGCCCGT CCTGATTCAT CCGGACTCGACCGTCTCGCC TTCTGCAACA 901 GTCGGACAAG GCGGCGTCGT TATGGCGAAA GCCGTCGTACAGGCTGACAG 951 CGTATTGAAA GACGGCGTAA TTGTGAACAC TGCCGCCACC GTCGATCACG1001 ATTGCCTGCT TGATGCTTTC GTCCACATCA GCCCGGGCGC GCACCTGTCG 1051GGCAACACGC GTATCGGCGA AGAAAGCTGG ATAGGCACAG GCGCGTGCAG 1101 CCGCCAGCAGATCCGTATCG GCAGCCGCGC AACCATTGGA GCGGGCGCAG 1151 TCGTCGTGCG CGACGTTTCAGACGGCATGA CCGTCGCGGG CAACCCGGCA 1201 AAACCATTGG CAGGCAAAAA TACCGAGACCCTGCGGTCGT AA

This is predicted to encode a protein having amino acid sequence <SEQ ID16>:

1 MSKFFKRLFD IVASASGLIF LSPVFLILIY LIRKNLGSPV FFFQERPGKD 51 GKPFKMVKFRSMHDALDSDG ILLPDGERLT PFGKKLRAAS LDELPELWNV 101 LKGDMSLVGP RPLLMQYLPLYDNFQNRRHE MKPGITGWAQ VNGRNALSWD 151 ERFACDIWYIDHFSLCLDIK ILLLTVKKVL IKEGISAQGE ATMPPFTGKR 201 KLAVVGAGGH GKVVAELAAALGTYGEIVFL DDRVQGSVNG FPVIGTTLLL 251 ENSLSPEQFD IAVAVGNNRI RRQIAEKAAALGFALPVLIH PDSTVSPSAT 301 VGQGGVVMAK AVVQADSVLK DGVIVNTAAT VDHDCLLDAFVHISPGAHLS 351 GNTRIGEESW IGTGACSRQQ IRIGSRATIG AGAVVVRDVS DGMTVAGNPA401 KPLAGKNTET LRS*

Two transmembrane domains are underlined.

ORF3-1 shows 94.6% identity in 410 aa overlap with ORF3a:

Homology with Hypothetical Protein Encoded by yvfc Gene (AccessionZ71928) of B. subtilis

ORF3 and YVFC proteins show 55% aa identity in 170 aa overlap (BLASTp):

ORF3 3 IYLIRKNLGSPVFFFQERPGKDGKPFKMVKFRSMRDGLYSDGIPLPDGERLTPFGKKLRA 62 I++R  +GSPVFF Q RPG  GKPF + KFR+M D   S G  LPD  RLT  G+ +R yvfc 27IAVVRLKIGSPVFFKQVRPGLHGKPFTLYKFRTMTDERDSKGNLLPDEVRLTKTGRLIRK 86 ORF3 63ASXDELPELWNILKGEMSLVGPRPLLMQYLPLYDNFQNRRHEMKPGITGWAQVNGRNALS 122  SDELP+L N+LKG++SLVGPRPLLM YLPLY   Q RRHE+KPGITGWAQ+NGRNA+S yvfc 87LSIDELPQLLNVLKGDLSLVGPRPLLMDYLPLYTEKQARRHEVKPGITGWAQINGRNAIS 146 ORF3123 WDEKFACDVWYIDHFSLCLDXXXXXXXXXXXXXXEGISAQGEXTMPPFTG 172W++KF  DVWY+D++S  LD              EGI      T   FTG yvfc 147WEKKFELDVWYVDNWSFFLDLKILCLTVRKVLVSEGIQQTNHVTAERFTG 196Homology with a Predicted ORF from N. gonorrhoeae

ORF3 shows 86.3% identity over a 286aa overlap with a predicted ORF(ORF3.ng) from N. gonorrhoeae:

The complete length ORF3ng nucleotide sequence <SEQ ID 17> is:

1 ATGAGTAAAG CCGTCAAACG CCTGTTCGAC ATCATCGCAT CCGCATCGGG 51 GCTGATTGTCCTGTCGCCCG TGTTTTTGGT TTTAATATAC CTCATCCGCA 101 AAAACTTAGG TTCGCCCGTCTTCTTCattC GGGAACGCCc cgGAAAGGAc 151 ggaaaacCTT TTAAAATGGT CAAATTCCGTTCCAtgcgcg acgcgcttGA 201 TTCAGACGGC ATTCCGCTGC CCGATAGCGA ACGCCTGACCGATTTCGGCA 251 AAAAATTACG CGCCACCAGT TTGGACGAAC TTCCTGAATT ATGGAATGTC301 CTCAAAGGCG AGATGAGCCT GGTCGGCCCC CGCCCGCTTT TGATGCAGTA 351TCTGCCGCTT TACAACAAAT TTCAAAACCG CCGCCACGAA ATGAAACCGG 401 GCATTACCGGCTGGGCGCAG GTCAACGGGC GCAACGCGCT TTCGTGGGAC 451 GAAAAGTTCT CCTGCGATGTTTGGTACACC GACAATTTCA GCTTTTGGCT 501 GGATATGAAA ATCCTGTTTC TGACAGTCAAAAAAGTCTTG ATTAAAGAAG 551 GCATTTCGGC GCAAGGGGAA GCCACCATGC CCCCTTTCGCGGGGAATCGC 601 AAACTCGCCG TTATCGGCGC GGGCGGACAC GGCAAAGTCG TTGCCGAGCT651 TGCCGCCGCA CTCGGCACAT ACGGCGAAAT CGTTTTTCTG GACGACCGCA 701CCCAAGGCAG CGTCAACGGC TTCCCCGTCA TCGGCACGAC GCTGCTGCTT 751 GAAAACAGTTTATCGCCCGA ACAATTCGAC ATCACCGTCG CCGTCGGCAA 801 CAACCGCATC CGCCGCCAAATCACCGAAAA CGCCGCCGCG CTCGGCTTCA 851 AACTGCCCGT TCTGATTCAT CCCGACGCGACCGTCTCGCC TTCTGCAATA 901 ATCGGACAAG GCAGCGTCGT AATGGCGAAA GCCGTCGTACAGGCCGGCAG 951 CGTATTGAAA GACGGCGTGA TTGTGAACAC TGCCGCCACC GTCGATCACG1001 ACTGCCTGCT TGACGCTTTC GtccaCATCA GCCCGGGCGC GCACCTGTCG 1051GGCAACACGC GTATCGGCGA AGAAAGCCGG ATAGGCACGG GCGCGTGCAG 1101 CCGCCAGCAGACAACCGTCG GCAGCGGGGT TACCgccgGT GCAGGGgcGG 1151 TTATCGTATG CGACATCCCGGACGGCATGA CCGTCGCGGG CAACCCGGCA 1201 AAGCCCCTTA CGGGCAAAAA CCCCAAGACCGGGACGGCAT AA

This encodes a protein having amino acid sequence <SEQ ID 18>:

1 MSKAVKRLFD IIASASGLIV LSPVFLVLIY LIRKNLGSPV FFIRERPGKD 51 GKPFKMVKFRSMRDALDSDG IPLPDSERLT DFGKKLRATS LDELPELWNV 101 LKGEMSLVGP RPLLMQYLPLYNKFQNRRHE MKPGITGWAQ VNGRNALSWD 151 EKFSCDVWYT DNFSFWLDMK ILFLTVKKVLIKEGISAQGE ATMPPFAGNR 201 KLAVIGAGGH GKVVAELAAA LGTYGEIVFL DDRTQGSVNGFPVIGTTLLL 251 ENSLSPEQFD ITVAVGNNRI RRQITENAAA LGFKLPVLIH PDATVSPSAI301 IGQGSVVMAK AVVQAGSVLK DGVIVNTAAT VDHDCLLDAF VHISPGAHLS 351GNTRIGEESR IGTGACSRQQ TTVGSGVTAG AGAVIVCDIP DGMTVAGNPA 401 KPLTGKNPKTGTA*

This protein shows 86.9% identity in 413 aa overlap with ORF3-1:

In addition, ORF3ng shows significant homology with a hypotheticalprotein from B. subtilis:

gnl|PID|e238668 (Z71928) hypothetical protein [Bacillussubtilis] >gi|1945702|gnl|PID|e313004 (Z94043) hypothetical protein[Bacillus subtilis] >gi|2635938|gnl|PID|e1186113 (Z99121) similar tocapsular polysaccharide biosynthesis [Bacillus subtilis] Length = 202Score = 235 bits (594), Expect = 3e−61 Identities = 114/195 (58%),Positives = 142/195 (72%) Query: 5VKRLFDIIASASGLIVLSPVFLVLIYLIRKNLGSPVFFIRERPGKDGKPFKMVKFRSMRD 64+KRLFD+ A+   L  S + L  I ++R  +GSPVFF + RPG  GKPF + KFR+M D Sbjct: 3LKRLFDLTAAIFLLCCTSVIILFTIAVVRLKIGSPVFFKQVRPGLHGKPFTLYKFRTMTD 62 Query:65 ALDSDGIPLPDSERLTDFGKKLRATSLDELPELWNVLKGEMSLVGPRPLLMQYLPLYNKF 124   DSG  LPD  RLT  G+ +R  S+DELP+L NVLKG++SLVGPRPLLM YLPLY + Sbjct: 63ERDSKGNLLPDEVRLTKTGRLIRKLSIDELPQLLNVLKGDLSLVGPRPLLMDYLPLYTEK 122 Query:125 QNRRHEMKPGITGWAQVNGRNALSWDEKFSCDVWYTDNFSFWLDMKILFLTVKKVLIKEG 184 QRRHE+KPGITGWAQ+NGRNA+SW++KF  DVWY DN+SF+LD+KIL LTV+KVL+ EG Sbjct: 123QARRHEVKPGITGWAQINGRNAISWEKKFELDVWYVDNWSFFLDLKILCLTVRKVLVSEG 182 Query:185 ISAQGEATMPPFAGN 199 I      T   F G+ Sbjct: 183 IQQTNHVTAERFTGS 197

The hypothetical product of yvfc gene shows similarity to EXOY of R.meliloti, an exopolysaccharide production protein. Based on this and onthe two predicted transmembrane regions in the homologous N. gonorrhoeaesequence, it is predicted that these proteins, or their epitopes, couldbe useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 4

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 19>:

1 ..AACCATATGG CGATTGTCAT CGACGAATAC GGCGGCACAT CCGGCTTGGT 51  CACCTTTGAA GACATCATCG AGCAAATCGT CGGCGAAATC GAAGACGAGT 101  TTGACGAAGA CGATAGCGCC GACAATATCC ATGCCGTTTC TTCAGACACG 151  TGGCGCATCC ATGCAGCTAC CGAAATCGAA GACATCAACA CCTTCTTCGG 201  CACGGAATAC AGCATCGAAG AAGCCGACAC CATT.GGCGG CCTGGTCATT 251  CAAGAGTTGG GACATCTGCC CGTGCGCGGC GAAAAAGTCC TTATCGGCGG 301  TTTGCAGTTC ACCGTCGCAC GCGCCGACAA CCGCCGCCTG CATACGCTGA 351  TGGCGACCCG CGTGAAGTAA GC........ .....ACCGC CGTTTCTGCA 401   CAGTTTAG

This corresponds to amino acid sequence <SEQ ID 20; ORF5>:

1 ..NHMAIVIDEY GGTSGLVTFE DIIEQIVGEI EDEFDEDDSA DNIHAVSSDT 51  WRIHAATEIE DINTFFGTEY SIEEADTIXR PGHSRVGTSA RARRKSPYRR 101  FAVHRRTRRQ PPPAYADGDP REVS....XR RFCTV*

Further sequence analysis revealed the complete DNA sequence to be <SEQID 21>:

1 ATGGACGGCG CACAACCGAA AACGAATTTT TTTGAACGCC TGATTGCCCG 51 ACTCGCCCGCGAACCCGATT CCGCCGAAGA CGTATTAAAC CTGCTTCGGC 101 AGGCGCACGA GCAGGAAGTTTTTGATGCGG ATACGCTTTT AAGATTGGAA 151 AAAGTCCTCG ATTTTTCCGA TTTGGAAGTGCGCGACGCGA TGATTACGCG 201 CAGCCGTATG AACGTTTTAA AAGAAAACGA CAGCATCGAGCGCATCACCG 251 CCTACGTTAT CGATACCGCC CATTCGCGCT TCCCCGTCAT CGGCGAAGAC301 AAAGACGAAG TTTTGGGCAT TTTGCACGCC AAAGACCTGC TCAAATATAT 351GTTTAACCCC GAGCAGTTCC ACCTCAAATC CATTCTCCGC CCCGCCGTCT 401 TCGTCCCCGAAGGCAAATCG CTGACCGCCC TTTTAAAAGA GTTCCGCGAA 451 CAGCGCAACC ATATGGCGATTGTCATCGAC GAATACGGCG GCACATCCGG 501 CTTGGTCACC TTTGAAGACA TCATCGAGCAAATCGTCGGC GAAATCGAAG 551 ACGAGTTTGA CGAAGACGAT AGCGCCGACA ATATCCATGCCGTTTCTTCC 601 GAACGCTGGC GCATCCATGC AGCTACCGAA ATCGAAGACA TCAACACCTT651 CTTCGGCACG GAATACAGCA GCGAAGAAGC CGACACCATT CGGCCTGGTC 701ATTCAAGAGT TGGGACATCT GCCCGTGCGC GGCGAAAAAG TCCTTATCGG 751 CGGTTTGCAGTTCACCGTCG CACGCGCCGA CAACCGCCGC CTGCATACGC 801 TGATGGCGAC CCGCGTGAAGTAAGCACCGC CGTTTCTGCA CAGTTTAGGA 851 TGACGGTACG GGCGTTTTCT GTTTCAATCCGCCCCATCCG CCAAACATAA

This corresponds to amino acid sequence <SEQ ID 22; ORF5-1>:

1 MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV FDADTLLRLE 51 KVLDFSDLEVRDAMITRSRM NVLKENDSIE RITAYVIDTA HSRFPVIGED 101 KDEVLGILHA KDLLKYMFNPEQFHLKSILR PAVFVPEGKS LTALLKEFRE 151 QRNHMAIVID EYGGTSGLVT FEDIIEQIVGEIEDEFDEDD SADNIHAVSS 201 ERWRIHAATE IEDINTFFGT EYSSEEADTI RPGHSRVGTSARARRKSPYR 251 RFAVHRRTRR QPPPAYADGD PREVSTAVSA QFRMTVRAFS VSIRPIRQT*

Further work identified the corresponding gene in strain A of N.meningitidis <SEQ ID 23>:

1 ATGGACGGCG CACAACCGAA AACAAATTTT TTNNAACGCC TGATTGCCCG 51 ACTCGCCCGCGAACCCGATT CCGCCGAAGA CGTATTGACC CTGTTGCGCC 101 AAGCGCACGA ACAGGAAGTATTTGATGCGG ATACGCTTTT AAGATTGGAA 151 AAAGTCCTCG ATTTTTCTGA TTTGGAAGTGCGCGACGCGA TGATTACGCG 201 CAGCCGTATG AACGTTTTAA AAGAAAACGA CAGCATCGAACGCATCACCG 251 CCTACGTTAT CGATACCGCC CATTCGCGCT TCCCCGTCAT CGGTGAAGAC301 AAAGACGAAG TTTTGGGTAT TTTGCACGCC AAAGACCTGC TCAAATATAT 351GTTCAACCCC GAGCAGTTCC ACCTCAAATC GATATTGCGC CCTGCCGTCT 401 TCGTCCCCGAAGGCAAATCG CTGACCGCCC TTTTAAAAGA GTTCCGCGAA 451 CAGCGCAACC ATATGGCAATCGTCATCGAC GAATACGGCG GCACGTCGGG 501 TTTGGTAACT TTTGAAGACA TCATCGAGCAAATCGTCGGC GACATCGAAG 551 ATGAGTTTGA CGAAGACGAA AGCGCGGACA ACATCCACGCCGTTTCCGCC 601 GAACGCTGGC GCATCCACGC GGCTACCGAA ATCGAAGACA TCAACGCCTT651 TTTCGGCACG GAATACAGCA GCGAAGAAGC CGACACCATC GGCGGCCNTG 701GTCATTCAGG AATTGGNACA CCTGCCCGTG CGCGGCGAAA AAGTCNTTAT 751 CGGCGNNTTGCANTTCACNG TCGCCNGCGC NGACAACCGC CGCCTGCATA 801 CGCTGATGGC GACCCGCGTGAAGTAAGCTC CGCCGTTTCT GTACAGTTTA 851 GGATGACGGT ACGGGCGTTT TCTGTTTCAATCCGCCCCAT CCGCCANACA 901 TAA

This encodes a protein having amino acid sequence <SEQ ID 24; ORF5a>:

1 MDGAQPKTNF XXRLIARLAR EPDSAEDVLT LLRQAHEQEV FDADTLLRLE 51 KVLDFSDLEVRDAMITRSRM NVLKENDSIE RITAYVIDTA HSRFPVIGED 101 KDEVLGILHA KDLLKYMFNPEQFHLKSILR PAVFVPEGKS LTALLKEFRE 151 QRNHMAIVID EYGGTSGLVT FEDIIEQIVGDIEDEFDEDE SADNIHAVSA 201 ERWRIHAATE IEDINAFFGT EYSSEEADTI GGXGHSGIGTPARARRKSXY 251 RRXAXHXRXR XQPPPAYADG DPREVSSAVS VQFRMTVRAF SVSIRPIRXT301 *

The originally-identified partial strain B sequence (ORF5) shows 54.7%identity over a 124aa overlap with ORF5a:

The complete strain B sequence (ORF5-1) and ORF5a show 92.7% identity in300 aa overlap:

Further work identified the a partial DNA sequence in N. gonorrhoeae<SEQ ID 25> which encodes a protein having amino acid sequence <SEQ ID26; ORF5ng>:

1 MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV FDADTLTRLE 51 KVLDFAELEVRDAMITRSRM NVLKENDSIE RITAYVIDTA HSRFPVIGED 101 KDEVLGILHA KDLLKYMFNPEQFHLKSVLR PAVFVPEGKS LTALLKEFRE 151 QRNHMAIVID EYGGTSGLVT FEDIIEQIVGDIEDEFDEDE SADDIHSVSA 201 ERWRIHAATE IEDINAFFGT EYGSEEADTI RRLGHSGIGTPARARRKSPY 251 RRFAVHRRPR RQPPPAHADG DPREVSRACP HRRFCTV*

Further analysis revealed the complete gonococcal nucleotide sequence<SEQ ID 27> to be:

1 ATGGACGGCG CACAACCGAA AACAAATTTT TTTGAACGCC TGATTGCCCG 51 ACTCGCCCGCGAACCCGATT CCGCCGAAGA CGTATTAAAC CTGCTTCGGC 101 AGGCGCACGA ACAGGAAGTTTTTGATGCCG ACACACTGAC CCGGCTGGAA 151 AAAGTATTGG ACTTTGCCGA GCTGGAAGTGCGCGATGCGA TGATTACGCG 201 CAGCCGCATG AACGTATTGA AAGAAAACGA CAGCATCGAACGCATCACCG 251 CCTACGTCAT CGATACCGCC CATTCGCGCT TCCCCGTCAT CGGCGAAGAC301 AAAGACGAAG TTTTGGGCAT TTTGCACGCC AAAGACCTGC TCAAATATAT 351GTTCAACCCC GAGCAGTTCC ACCTGAAATC CGTCTTGCGC CCTGCCGTTT 401 TCGTGCCCGAAGGCAAATCT TTGACCGCCC TTTTAAAAGA GTTCCGCGAA 451 CAGCGCAACC ATATGGCAATCGTCATCGAC GAATACGGCG GCACGTCGGG 501 TTTGGTCACC TTTGAAGACA TCATCGAGCAAATCGTCGGT GACATCGAAG 551 ACGAGTTTGA CGAAGACGAA AGCGccgacg acatCCACTCcgTTTccgCC 601 GAACGCTGGC GCATCCacgc ggctaCCGAA ATCGAAGaca TCAACGCCTT651 TTTCGGTACG GAatacggca gcgaagaagc cgacaccatc cggcggctTG 701GTCATTCAGG AATTGGGACA CCTGCCCGTG CGCGGCGAAA AAGTCCTTAt 751 cggcgGTTTGCagttcaccg tCGCCCGCGC CGACAACCGC CGCCTGCACA 801 CGCTGATGGC GACCCGCGTGAAGTAAGCAG AGCCTGCCcg AccgccgttT 851 CTGCacAGTT TAGGatgACG gtaCGGTCGTTTTCTGTTTC AATCCGCCCC 901 ATCCGCCAAA CATAA

This encodes a protein having amino acid sequence <SEQ ID 28; ORF5ng-1>:

1 MDGAQPKTNF FERLIARLAR EPDSAEDVLN LLRQAHEQEV FDADTLTRLE 51 KVLDFAELEVRDAMITRSRM NVLKENDSIE RITAYVIDTA HSRFPVIGED 101 KDEVLGILHA KDLLKYMFNPEQFHLKSVLR PAVFVPEGKS LTALLKEFRE 151 QRNHMAIVID EYGGTSGLVT FEDIIEQIVGDIEDEFDEDE SADDIHSVSA 201 ERWRIHAATE IEDINAFFGT EYGSEEADTI RRLGHSGIGTPARARRKSPY 251 RRFAVHRRPR RQPPPAHADG DPREVSRACP TAVSAQFRMT VRSFSVSIRP301 IRQT*

The originally-identified partial strain B sequence (ORFS) shows 83.1%identity over a 135aa overlap with the partial gonococcal sequence(ORF5ng):

The complete strain B and gonococcal sequences (ORFS-1 & ORF5ng-1) show92.4% identity in 304 aa overlap:

Computer analysis of these amino acid sequences indicates a putativeleader sequence, and identified the following homologies:

Homology with Hemolysin Homolog TlyC (Accession U32716) of H. influenzae

ORF5 and TlyC proteins show 58% aa identity in 77 aa overlap (BLASTp).

ORF5 2 HMAIVIDEYGGTSGLVTFEDIIEQIVGEIEDEFDEDDSADNIHAVSSDTWRIHAATEIED 61HMAIV+DE+G  SGLVT EDI+EQIVG+IEDEFDE++ AD I  +S  T+ + A T+I+D TlyC 166HMAIVVDEFGAVSGLVTIEDILEQIVGDIEDEFDEEEIAD-IRQLSRHTYAVRALTDIDD 224 ORF5 62INTFFGTEYSIEEADTI 78  N  F T++  EE DTI TlyC 225 FNAQFNTDFDDEEVDTI 241

ORF5ng-1 also shows significant homology with TlyC:

-   -   SCORES Init1: 301 Initn: 419 Opt: 668    -   Smith-Waterman score: 668; 45.9% identity in 242 aa overlap

Homology with a Hypothetical Secreted Protein from E. coli:

ORF5a shows homology to a hypothetical secreted protein from E. coli:

sp|P77392|YBEX_ECOLI HYPOTHETICAL 33.3 KD PROTEIN IN CUTE-ASNBINTERGENIC REGION >gi|1778577 (U82598) similar to H. influenzae[Escherichia coli] >gi|1786879 (AE000170) f292; This 292 aa ORF is 23%identical (9 gaps) to 272 residues of an approx. 440 aa proteinYTFL_HAEIN SW: P44717 [Escherichia coli] Length = 292 Score = 212 bits(533), Expect = 3e−54 Identities = 112/230 (48%), Positives = 149/230(64%), Gaps = 3/230 (1%) Query: 2DGAQPKTNFXXRLIARLAR-EPDSAEDVLTLLRQAHEQEVFDADTLLRLEKVLDFSDLEV 60D    K  F   L+++L   EP + +++L L+R + + ++ D DT   LE V+D +D  V Sbjct: 10DTISNKKGFFSLLLSQLFHGEPKNRDELLALIRDSGQNDLIDEDTRDMLEGVMDIADQRV 69 Query:61 RDAMITRSRMNVLKENDSIERITAYVIDTAHSRFPVIGEDKDEVLGILHAKDLLKYM-FN 119 RDMI RS+M  LK N +++     +I++AHSRFPVI EDKD + GIL AKDLL +M  + Sbjct: 70RDIMIPRSQMITLKRNQTLDECLDVIIESAHSRFPVISEDKDHIEGILMAKDLLPFMRSD 129 Query:120 PEQFHLKSILRPAVFVPEGKSLTALLKEFREQRNHMAIVIDEYGGTSGLVTFEDIIEQIV 179  EF +  +LR AV VPE K +  +LKEFR QR HMAIVIDE+GG SGLVT EDI+E IV Sbjct: 130AEAFSMDKVLRQAVVVPESKRVDRMLKEFRSQRYHMAIVIDEFGGVSGLVTIEDILELIV 189 Query:180 GDIEDEFDEDESADNIHAVSAERWRIHAATEIEDINAFFGTEYSSEEADT 229G+IEDE+DE++  D    +S   W + A   IED N  FGT +S EE DT Sbjct: 190GEIEDEYDEEDDID-FRQLSRHTWTVRALASIEDFNEAFGTHFSDEEVDT 238

Based on this analysis, including the amino acid homology to the TlyChemolysin-homologue from H. influenzae (hemolysins are secretedproteins), it was predicted that the proteins from N. meningitidis andN. gonorrhoeae are secreted and could thus be useful antigens forvaccines or diagnostics.

ORF5-1 (30.7 kDa) was cloned in the pGex vector and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 2A shows the results ofaffinity purification of the GST-fusion protein. Purified GST-fusionprotein was used to immunise mice, whose sera were used for Western blotanalysis (FIG. 1B). These experiments confirm that ORFS-1 is asurface-exposed protein, and that it is a useful immunogen.

Example 5

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 29>:

1 ATGCGCGGCG GCAGGCCGGA TTCCGTTACC GTGCAGATTA TCGAAGGTTC 51 GCGTTTTTCGCATATGAGGA AAGTCATCGA CGCAACGCCC GACATCGGAC 101 ACGACACCAA AGGCTGGAGCAATGAAAAAC TGATGGCGGA AGTTGCGCCC 151 GATGCCTTCA GCGGCAATCC TGAAgGGCAGTTTTTCCCCG ACAGCTACGA 201 AATCGATGCG GGCGGCAGTG ATTTGCAGAT TTACCAAACCGCCTACAAgG 251 GCGATGCAAC GCCGCCTGAA TGAgGGCATG GGAAAGCAGG CAGGACGGGC301 TGCCTTATAA AAACCCTTAT GAAATGCTGA TTATGGCGAr CCTGGTCGAA 351AAGGAAACAG GGCATGAAGC CGAsCsCGAC CATGTcGCTT CCGTCTTCGT 401 CAACCGCCTGAAAATCGGTA TGCGCCTGCA AACCgAssCG TCCGTGATTT 451 ACGGCATGGG TGCGGCATACAAGGGCAAAA TCCGTAAAGC CGACCTGCGC 501 CGCGACACGC CGTACAACAC CTACACGCGCGGCGGTCTGC CGCCAACCCC 551 GATTGCGCTG CCC..

This corresponds to the amino acid sequence <SEQ ID 30; ORF7>:

1 MRGGRPDSVT VQIIEGSRFS HMRKVIDATP DIGHDTKGWS NEKLMAEVAP 51 DAFSGNPEGQFFPDSYEIDA GGSDLQIYQT AYKAMQRRLN EAWESRQDGL 101 PYKNPYEMLI MAXLVEKETGHEAXXDHVAS VFVNRLKIGM RLQTXXSVIY 151 GMGAAYKGKI RKADLRRDTP YNTYTRGGLPPTPIALP..

Further sequence analysis revealed the complete DNA sequence <SEQ ID31>:

1 ATGTTGAGAA AATTGTTGAA ATGGTCTGCC GTTTTTTTGA CCGTGTCGGC 51 AGCCGTTTTCGCCGCGCTGC TTTTTGTTCC TAAGGATAAC GGCAGGGCAT 101 ACCGAATCAA AATTGCCAAAAACCAGGGTA TTTCGTCGGT CGGCAGGAAA 151 CTTGCCGAAG ACCGCATCGT GTTCAGCAGGCATGTTTTGA CGGCGGCGGC 201 CTACGTTTTG GGTGTGCACA ACAGGCTGCA TACGGGGACGTACAGATTGC 251 CTTCGGAAGT GTCTGCTTGG GATATCTTGC AGAAAATGCG CGGCGGCAGG301 CCGGATTCCG TTACCGTGCA GATTATCGAA GGTTCGCGTT TTTCGCATAT 351GAGGAAAGTC ATCGACGCAA CGCCCGACAT CGGACACGAC ACCAAAGGCT 401 GGAGCAATGAAAAACTGATG GCGGAAGTTG CGCCCGATGC CTTCAGCGGC 451 AATCCTGAAG GGCAGTTTTTCCCCGACAGC TACGAAATCG ATGCGGGCGG 501 CAGTGATTTG CAGATTTACC AAACCGCCTACAAGGCGATG CAACGCCGCC 551 TGAATGAGGC ATGGGAAAGC AGGCAGGACG GGCTGCCTTATAAAAACCCT 601 TATGAAATGC TGATTATGGC GAGCCTGGTC GAAAAGGAAA CAGGGCATGA651 AGCCGACCGC GACCATGTCG CTTCCGTCTT CGTCAACCGC CTGAAAATCG 701GTATGCGCCT GCAAACCGAC CCGTCCGTGA TTTACGGCAT GGGTGCGGCA 751 TACAAGGGCAAAATCCGTAA AGCCGACCTG CGCCGCGACA CGCCGTACAA 801 CACCTACACG CGCGGCGGTCTGCCGCCAAC CCCGATTGCG CTGCCCGGCA 851 AGGCGGCACT CGATGCCGCC GCCCATCCGTCCGGCGAAAA ATACCTGTAT 901 TTCGTGTCCA AAATGGACGG CACGGGCTTG AGCCAGTTCAGCCATGATTT 951 GACCGAACAC AATGCCGCCG TCCGCAAATA TATTTTGAAA AAATAA

This corresponds to the amino acid sequence <SEQ ID 32; ORF7-1>:

1 MLRKLLKWSA VFLTVSAAVF AALLFVPKDN GRAYRIKIAK NQGISSVGRK 51 LAEDRIVFSRHVLTAAAYVL GVHNRLHTGT YRLPSEVSAW DILQKMRGGR 101 PDSVTVQIIE GSRFSHMRKVIDATPDIGHD TKGWSNEKLM AEVAPDAFSG 151 NPEGQFFPDS YEIDAGGSDL QIYQTAYKAMQRRLNEAWES RQDGLPYKNP 201 YEMLIMASLV EKETGHEADR DHVASVFVNR LKIGMRLQTDPSVIYGMGAA 251 YKGKIRKADL RRDTPYNTYT RGGLPPTPIA LPGKAALDAA AHPSGEKYLY301 FVSKMDGTGL SQFSHDLTEH NAAVRKYILK K*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical Protein Encoded by yceg Gene (AccessionP44270) of H. influenzae

ORF7 and yceg proteins show 44% aa identity in 192 aa overlap:

ORF7   1 MRGGRPDSVTVQIIEGSRFSHMRKVIDATPDIGHDTKGWSNEKLMA-----EVAPDAFSG  55         +  G+     V+ IEG  F   RK ++  P +    K  SNE++ A      ++  +yceg 102 LNSGKEVQFNVKWIEGKTFKDWRKDLENAPHLVQTLKDKSNEEIFALLDLPDIGQNLELK 161ORF7  56 NPEGQFFPDSYEIDAGGSDLQIYQTAYKAMQRRLNEAWESRQDGLPYKNPYEMLIMAXLV 115         N EG  +PD+Y      +DL++ + + + M++ LN+AW  R + LP   NPYEMLI+A +Vyceg 162 NVEGWLYPDTYNYTPKSTDLELLKRSAERMKKALNKAWNERDEDLPLANPYEMLILASIV 221ORF7 116 EKETGHEAXXDHVASVFVNRLKIGMRLQTXXSVIYGMGAAYKGKIRKADLRRDTPYNTYT 175         EKETG       VASVF+NRLK  M+LQT  +VIYGMG  Y G IRK DL   TPYNTYyceg 222 EKETGIANERAKVASVFINRLKAKMKLQTDPTVIYGMGENYNGNIRKKDLETKTPYNTYV 281ORF7 176 RGGLPPTPIALP                                                 187           GLPPTPIA+Pyceg 282 IDGLPPTPIAMP                                                 293

The complete length YCEG protein has sequence:

1 MKKFLIAILL LILILAGVAS FSYYKMTEFV KTPVNVQADE LLTIERGTTS 51 SKLATLFEQEKLIADGKLLP YLLKLKPELN KIKAGTYSLE NVKTVQDLLD 101 LLNSGKEVQF NVKWIEGKTFKDWRKDLENA PHLVQTLKDK SNEEIFALLD 151 LPDIGQNLEL KNVEGWLYPD TYNYTPKSTDLELLKRSAER MKKALNKAWN 201 ERDEDLPLAN PYEMLILASI VEKETGIANE RAKVASVFINRLKAKMKLQT 251 DPTVIYGMGE NYNGNIRKKD LETKTPYNTY VIDGLPPTPI AMPSESSLQA301 VANPEKTDFY YFVADGSGGH KFTRNLNEHN KAVQEYLRWY RSQKNAKHomology with a Predicted ORF from N. meningitidis (Strain A)

ORF7 shows 95.2% identity over a 187aa overlap with an ORF (ORF7a) fromstrain A of N. meningitidis:

The complete length ORF7a nucleotide sequence <SEQ ID 33> is:

1 ATGTTGAGAA AATTGTTGAA ATGGTCTGCC GTTTTTTTGA CCGTATCGGC 51 AGCCGTTTTCGCCGCGCTGC TTTTCGTCCC TAAAGACAAC GGCAGGGCAT 101 ACAGGATTAA AATTGCCAAAAACCAGGGTA TTTCGTCGGT CGGCAGGAAA 151 CTTGCCGAAG ACCGCATCGT GTTCAGCAGGCATGTTTTGA CGGCGGCGGC 201 CTACGTTTTG GGTGTGCACA ACAGGCTGCA TACGGGGACGTACAGACTGC 251 CTTCGGAAGT GTCTGCTTGG GATATCTTGC AGAAAATGCG CGGCGGCAGG301 CCGGATTCCG TTACCGTGCA GATTATCGAA GGTTCGCGTT TTTCGCATAT 351GAGGAAAGTC ATCGACGCAA CGCCCGACAT CGAACACGAC ACCAAAGGCT 401 GGAGCAATGAAAAACTGATG GCGGAAGTTG CCCCTGATGC CTTCAGCGGC 451 AATCCTGAAG GGCAGTTTTTCCCCGACAGC TACGAAATCG ATGCGGGCGG 501 CAGCGATTTA CGGATTTACC AAATCGCCTACAAGGCGATG CAACGCCGAC 551 TGAATGAGGC ATGGGAAAGC AGGCAGGACG GGCTGCCTTATAAAAACCCT 601 TATGAAATGC TGATTATGGC GAGCCTGATC GAAAAGGAAA CAGGGCATGA651 AGCCGACCGC GACCATGTCG CTTCCGTCTT CGTCAACCGC CTGAAAATCG 701GTATGCGCCT GCAAACCGAC CCGTCCGTGA TTTACGGCAT GGGTGCGGCA 751 TACAAGGGCAAAATCCGTAA AGCCGACCTG CGCCGCGACA CGCCGTACAA 801 CACCTACACG CGCGGCGGTCTGCCGCCAAC CCCGATCGCG CTGCCCGGCA 851 AGGCGGCACT CGATGCCGCC GCCCATCCGTCCGGTGAAAA ATACCTGTAT 901 TTCGTGTCCA AAATGGACGG TACGGGCTTG AGCCAGTTCAGCCATGATTT 951 GACCGAACAC AACGCCGCCG TTCGCAAATA TATTTTGAAA AAATAA

This is predicted to encode a protein having amino acid sequence <SEQ ID34>:

1 MLRKLLKWSA VFLTVSAAVF AALLFVPKDN GRAYRIKIAK NQGISSVGRK 51 LAEDRIVFSRHVLTAAAYVL GVHNRLHTGT YRLPSEVSAW DILQKMRGGR 101 PDSVTVQIIE GSRFSHMRKVIDATPDIEHD TKGWSNEKLM AEVAPDAFSG 151 NPEGQFFPDS YEIDAGGSDL RIYQIAYKAMQRRLNEAWES RQDGLPYKNP 201 YEMLIMASLI EKETGHEADR DHVASVFVNR LKIGMRLQTDPSVIYGMGAA 251 YKGKIRKADL RRDTPYNTYT RGGLPPTPIA LPGKAALDAA AHPSGEKYLY301 FVSKMDGTGL SQFSHDLTEH NAAVRKYILK K*

A leader peptide is underlined.

ORF7a and ORF7-1 show 98.8% identity in 331 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF7 shows 94.7% identity over a 187aa overlap with a predicted ORF(ORF7.ng) from N. gonorrhoeae:

An ORF7ng nucleotide sequence <SEQ ID 35> is predicted to encode aprotein having amino acid sequence <SEQ ID 36>:

1 MRGGRPDSVT VQIIEGSRFS HMRKVIDATP DIGHDTKGWS NEKLMAEVAP 51 DAFSGNPEGQFFPDSYEIDA GGSDLQIYQT AYKAMQRRLN EAWAGRQDGL 101 PYKNPYEMLI MASLIEKETGHEADRDHVAS VFVNRLKIGM RLQTDPSVIY 151 GMGAAYKGKI RKADLRRDTP YNTYTGGGLPPTRIALPGKA AMDAAAHPSG 201 EKYLYFVSKM DGTGLSQFSH DLTEHNAAVR KYILKK*

Further sequence analysis revealed a partial DNA sequence of ORF7ng <SEQID 37>:

1 ..taccgaatca AGATTGCCAA AAATCAGGGT ATTTCGTCGG TCGGCAGGAA 51  ACTTGCcgaA GACCGCATCG TGTTCAGCAG GCATGTTTTG ACAGCGGCGG 101  CCTACGTTTT GGGTGTGCAC AACAGGCTGC ATACGGGGAC gTACAGATTG 151  CCTTCGGAAG TGTCTGCTTG GGATATCTTG CAGAAAATGC GCGGCGGCAG 201  GCCGGATTCC GTTACCGTGC AGATTATCGA AGGTTCGCGT TTTTCGCATA 251  TGAGGAAAGT CATCGACGCA ACGCCCGACA TCGGACACGA CACCAAAGGC 301  TGGAGCAATG AAAAACTGAT GGCGGAAGTT GCGCCCGATG CCTTCAGCGG 351  CAATCCTGAA GGGCAGTTTT TTCCCGACAG CTACGAAATC GATGCGGGCG 401  GCAGCGATTT GCAGATTTAC CAAACCGCCT ACAAGGCGAT GCAACGCCGC 451  CTGAACGAGG CATGGGCAGG CAGGCAGGAC GGGCTGCCTT ATAAAAACCC 501  TTATGAAATG CTGATTATGG CGAGCCTGAT CGAAAAGGAA ACGGGGCATG 551  AGGCCGACCG CGACCATGTC GCTTCCGTCT TCGTCAACCG CCTGAAAATC 601  GGTATGCGCC TGCAAACCGA CCCGTCCGTG ATTTACGGCA TGGGTGCGGC 651  ATACAAGGGC AAAATCCGTA AAGCCGACCT GCGCCGCGAC ACGCCGTACA 701  aCAccTAtac gggcgggggc ttgccgccaa cccggattgc gctgcccggC 751  Aaggcggcaa tggatgccgc cgcccacccg tccggcgaAa aatacctgTa 801  tttcgtgtcC AAAATGGACG GCACGGGCTT GAGCCAGTTC AGCCATGATT 851  TGACCGAACA CAACGCCGCc gTcCGCAAAT ATATTTTGAA AAAATAA

This corresponds to the amino acid sequence <SEQ ID 38; ORF7ng-1>:

1 ..YRIKIAKNQG ISSVGRKLAE DRIVFSRHVL TAAAYVLGVH NRLHTGTYRL 51  PSEVSAWDIL QKMRGGRPDS VTVQIIEGSR FSHMRKVIDA TPDIGHDTKG 101  WSNEKLMAEV APDAFSGNPE GQFFPDSYEI DAGGSDLQIY QTAYKAMQRR 151  LNEAWAGRQD GLPYKNPYEM LIMASLIEKE TGHEADRDHV ASVFVNALKI 201  GMRLQTDPSV IYGMGAAYKG KIRKADLRRD TPYNTYTGGG LPPTRIALPG 251  KAAMDAAAHP SGEKYLYFVS KMDGTGLSQF SHDLTEHNAA VRKYILKK*

ORF7ng-1 and ORF7-1 show 98.0% identity in 298 aa overlap:

In addition, ORF7ng-1 shows significant homology with a hypothetical E.coli protein:

sp|P28306|YCEG_ECOLI HYPOTHETICAL 38.2 KD PROTEIN IN PABC-HOLBINTERGENIC REGION gi|1787339 (AE000210) o340; 100% identical to fragmentYCEG_ECOLI SW: P28306 but has 97 additional C-terminal residues[Escherichia coli] Length = 340 Score = 79 (36.2 bits), Expect= 5.0e−57, Sum P(2) = 5.0e−57 Identities = 20/87 (22%), Positives= 40/87 (45%) Query: 10GISSVGRKLAEDRIVFSRHVLTAAAYVLGVHNRLHTGTYRLPSEVSAWDILQKMRGGRPD 69 G  ++G+L  D+I+    V      +    +    GTYR   +++  ++L+ +  G+ Sbjct: 49GRLALGEQLYADKIINRPRVFQWLLRIEPDLSHFKAGTYRFTPQMTVREMLKLLESGKEA 108 Query:70 SVTVQIIEGSRFSHMRKVIDATPDIGH 96    ++++EG R S   K +   P I H Sbjct: 109QFPLRLVEGMRLSDYLKQLREAPYIKH 135 Score = 438 (200.7 bits), Expect= 5.0e−57, Sum P(2) = 5.0e−57 Identities = 84/155 (54%), Positives= 111/155 (71%) Query: 120EGQFFPDSYEIDAGGSDLQIYQTAYKAMQRRLNEAWAGRQDGLPYKNPYEMLIMASLIEK 179 EGF+PD++   A  +D+ + + A+K M + ++ AW GR DGLPYK+  +++ MAS+IEK Sbjct: 158EGWFWPDTWMYTANTTDVALLKRAHKKMVKAVDSAWEGRADGLPYKDKNQLVTMASIIEK 217 Query:180 ETGHEADRDHVASVFVNRLKIGMRLQTDPSVIYGMGAAYKGKIRKADLRRDTPYNTYTGG 239ET   ++RD VASVF+NRL+IGMRLQTDP+VIYGMG  Y GK+ +ADL   T YNTYT Sbjct: 218ETAVASERDKVASVFINRLRIGMRLQTDPTVIYGMGERYNGKLSRADLETPTAYNTYTIT 277 Query:240 GLPPTRIALPGKAAMDAAAHPSGEKYLYFVSKMDG 274 GLPP  IAPG  ++ AAAHP+   YLYFV+   G Sbjct: 278GLPPGAIATPGADSLKAAAHPAKTPYLYFVADGKG 312

Based on this analysis, including the fact that the H. influenzae YCEGprotein possesses a possible leader sequence, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 6

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 39>:

1 CGTTTCAAAA TGTTAACTGT GTTGACGGCA ACCTTGATTG CCGGACAGGT 51 ATCTGCCGCCGGAGGCGGTG CGGGGGATAT GAAACAGCCG AAGGAAGTCG 101 GAAAGGTTTT CAGAAAGCAGCAGCGTTACA GCGAGGAAGA AATCAAAAAC 151 GAACGCGCAC GGCTTGCGGC AGTGGGCGAGCGGGTTAATC AGATATTTAC 201 GTTGCTGGGA GGGGAAACCG CCTTGCAAAA GGGGCAGGCGGGAACGGCTC 251 TGGCAACCTA TATGCTGATG TTGGAACGCA CAAAATCCCC CGAAGTCGCC301 GAACGCGCCT TGGAAATGGC CGTGTCGCTG AACGCGTTTG AACAGGCGGA 351AATGATTTAT CAGAAATGGC GGCAGATTGA GCCTATACCG GGTAAGGCGC 401 AAAAACGGGCGGGGTGGCTG CGGAACGTGC TGAGGGAAAG AGGAAATCAG 451 CATCTGGACG GACGGGAAGAAGTGCTGGCT CAGGCGGACG AAGGACAG

This corresponds to the amino acid sequence <SEQ ID 40; ORF9>:

1 ..RFKMLTVLTA TLIAGQVSAA GGGAGDMKQP KEVGKVFRKQ QRYSEEEIKN 51  ERARLAAVGE RVNQIFTLLG GETALQKGQA GTALATYMLM LERTKSPEVA 101  ERALEMAVSL NAFEQAEMIY QKWRQIEPIP GKAQKRAGWL RNVLRERGNQ 151  HLDGREEVLA QADEGQ

Further sequence analysis revealed the complete DNA sequence <SEQ ID41>:

1 ATGTTACCTA ACCGTTTCAA AATGTTAACT GTGTTGACGG CAACCTTGAT 51 TGCCGGACAGGTATCTGCCG CCGGAGGCGG TGCGGGGGAT ATGAAACAGC 101 CGAAGGAAGT CGGAAAGGTTTTCAGAAAGC AGCAGCGTTA CAGCGAGGAA 151 GAAATCAAAA ACGAACGCGC ACGGCTTGCGGCAGTGGGCG AGCGGGTTAA 201 TCAGATATTT ACGTTGCTGG GAGGGGAAAC CGCCTTGCAAAAGGGGCAGG 251 CGGGAACGGC TCTGGCAACC TATATGCTGA TGTTGGAACG CACAAAATCC301 CCCGAAGTCG CCGAACGCGC CTTGGAAATG GCCGTGTCGC TGAACGCGTT 351TGAACAGGCG GAAATGATTT ATCAGAAATG GCGGCAGATT GAGCCTATAC 101 CGGGTAAGGCGCAAAAACGG GCGGGGTGGC TGCGGAACGT GCTGAGGGAA 451 AGAGGAAATC AGCATCTGGACGGACTGGAA GAAGTGCTGG CTCAGGCGGA 501 CGAAGGACAG AACCGCAGGG TGTTTTTATTGTTGGCACAA GCCGCCGTGC 551 AACAGGACGG GTTGGCGCAA AAAGCATCGA AAGCGGTTCGCCGCGCGGCG 601 TTGAAATATG AACATCTGCC CGAAGCGGCG GTTGCCGATG TGGTGTTCAG651 CGTACAGGGA CGCGAAAAGG AAAAGGCAAT CGGAGCTTTG CAGCGTTTGG 701CGAAGCTCGA TACGGAAATA TTGCCCCCCA CTTTAATGAC GTTGCGTCTG 751 ACTGCACGCAAATATCCCGA AATACTCGAC GGCTTTTTCG AGCAGACAGA 801 CACCCAAAAC CTTTCGGCCGTCTGGCAGGA AATGGAAATT ATGAATCTGG 851 TTTCCCTGCA CAGGCTGGAT GATGCCTATGCGCGTTTGAA CGTGCTGTTG 901 GAACGCAATC CGAATGCAGA CCTGTATATT CAGGCAGCGATATTGGCGGC 951 AAACCGAAAA GAAGGTGCTT CCGTTATCGA CGGCTACGCC GAAAAGGCAT1001 ACGGCAGGGG GACGGAGGAA CAGCGGAGCA GGGCGGCGCT AACGGCGGCG 1051ATGATGTATG CCGACCGCAG GGATTACGCC AAAGTCAGGC AGTGGCTGAA 1101 AAAAGTATCCGCGCCGGAAT ACCTGTTCGA CAAAGGTGTG CTGGCGGCTG 1151 CGGCGGCTGT CGAGTTGGACGGCGGCAGGG CGGCTTTGCG GCAGATCGGC 1201 AGGGTGCGGA AACTTCCCGA ACAGCAGGGGCGGTATTTTA CGGCAGACAA 1251 TTTGTCCAAA ATACAGATGC TCGCCCTGTC GAAGCTGCCCGATAAACGGG 1301 AGGCTTTGAG GGGGTTGGAC AAGATTATCG AAAAACCGCC TGCCGGCAGT1351 AATACAGAGT TACAGGCAGA GGCATTGGTA CAGCGGTCAG TTGTTTACGA 1401TCGGCTTGGC AAGCGGAAAA AAATGATTTC AGATCTTGAA AGGGCGTTCA 1451 GGCTTGCACCCGATAACGCT CAGATTATGA ATAATCTGGG CTACAGCCTG 1501 CTGACCGATT CCAAACGTTTGGACGAAGGT TTCGCCCTGC TTCAGACGGC 1551 ATACCAAATC AACCCGGACG ATACCGCTGTCAACGACAGC ATAGGCTGGG 1601 CGTATTACCT GAAAGGCGAC GCGGAAAGCG CGCTGCCGTATCTGCGGTAT 1651 TCGTTTGAAA ACGACCCCGA GCCCGAAGTT GCCGCCCATT TGGGCGAAGT1701 GTTGTGGGCA TTGGGCGAAC GCGATCAGGC GGTTGACGTA TGGACGCAGG 1751CGGCACACCT TACGGGAGAC AAGAAAATAT GGCGGGAAAC GCTCAAACGT 1801 CACGGCATCGCATTGCCCCA ACCTTCCCGA AAACCTCGGA AATAA

This corresponds to the amino acid sequence <SEQ ID 42; ORF9-1>:

1 MLPNRFKMLT VLTATLIAGQ VSAAGGGAGD MKQPKEVGKV FRKQQRYSEE 51 EIKNERARLAAVGERVNQIF TLLGGETALQ KGQAGTALAT YMLMLERTKS 101 PEVAERALEM AVSLNAFEQAEMIYQKWRQI EPIPGKAQKR AGWLRNVLRE 151 RGNQHLDGLE EVLAQADEGQ NRRVFLLLAQAAVQQDGLAQ KASKAVRRAA 201 LKYEHLPEAA VADVVFSVQG REKEKAIGAL QRLAKLDTEILPPTLMTLRL 251 TARKYPEILD GFFEQTDTQN LSAVWQEMEI MNLVSLHRLD DAYARLNVLL301 ERNPNADLYI QAAILAANRK EGASVIDGYA EKAYGRGTEE QRSRAALTAA 351MMYADRRDYA KVRQWLKKVS APEYLFDKGV LAAAAAVELD GGRAALRQIG 401 RVRKLPEQQGRYFTADNLSK IQMLALSKLP DKREALRGLD KIIEKPPAGS 451 NTELQAEALV QRSVVYDRLGKRKKMISDLE RAFRLAPDNA QIMNNLGYSL 501 LTDSKRLDEG FALLQTAYQI NPDDTAVNDSIGWAYYLKGD AESALPYLRY 551 SFENDPEPEV AAHLGEVLWA LGERDQAVDV WTQAAHLTGDKKIWRETLKR 601 HGIALPQPSR KPRK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF9 shows 89.8% identity over a 166aa overlap with an ORF (ORF9a) fromstrain A of N. meningitidis:

The complete length ORF9a nucleotide sequence <SEQ ID 43> is:

1 ATGTTACCCG CCCGTTTCAC CATTTTATCT GTGCTCGCGG CAGCCCTGCT 51 TGCCGGGCAGGCGTATGCCG CCGGCGCGGC GGATGCGAAG CCGCCGAAGG 101 AAGTCGGAAA GGTTTTCAGAAAGCAGCAGC GTTACAGCGA GGAAGAAATC 151 AAAAACGAAC GCGCACGGCT TGCGGCAGTGGGCGAGCGGG TTAATCAGAT 201 ATTTACGTTG CTGGGANGGG AAACCGCCTT GCAAAAGGGGCAGGCGGGAA 251 CGGCTCTGGC AACCTATATG CTGATGTTGG AACGCACAAA ATCCCCCGAA301 GTCGCCGAAC GCGCCTTGGA AATGGCCGTG TCNCTGAACG CGTTTGAACA 351GGCGGAAATG ATTTATCAGA AATGGCGGCA GATTGAGCCT ATACCGGGTA 401 AGGCGCAAAAACGGGCGGGG TGGCTGCGGA ACGTGCTGAG GGAAAGAGGA 451 AATCAGCATC TAGACGGACTGGAAGAANTG CTGGCTCAGG CGGACGAANG 501 ACAGAACCGC AGGGTGTTTT TATTGTTGGCACAAGCCGCC GTGCAACAGG 551 ACGGGTTGGC GCAAAAAGCA TCGAAAGCGG TTCGCCGCGCGGCGTTGAGA 601 TATGAACATC TGCCCGAAGC GGCGGTTGCC GATGTGGTGT TCAGCGTACA651 GGNACGCGAA AAGGAAAAGG CAATCGGAGC TTTGCAGCGT TTGGCGAAGC 701TCGATACGGA AATATTGCCC CCCACTTTAA TGACGTTGCG TCTGACTGCA 751 CGCAAATATCCCGAAATACT CGACGGCTTT TTCGAGCAGA CAGACACCCA 801 AAACCTTTCG GCCGTCTGGCAGGAAATGGA AATTATGAAT CTGGTTTCCC 851 TGCACAGGCT GGATGATGCC TATGCGCGTTTGAACGTGCT GTTGGAACGC 901 AATCCGAATG CAGACCTGTA TATTCAGGCA GCGATATTGGCGGCAAACCG 951 AAAAGAANGT GCTTCCGTTA TCGACGGCTA CGCCGAAAAG GCATACGGCA1001 GGGGGACGGG GGAACAGCGG GGCAGGGCGG CAATGACGGC GGCGATGATA 1051TATGCCGACC GAAGGGATTA CACCAAAGTC AGGCAGTGGT TGAAAAAAGT 1101 GTCCGCGCCGGAATACCTGT TCGACAAAGG TGTGCTGGCG GCTGCGGCGG 1151 CTGTCGAGTT GGACNGCGGCAGGGCGGCTT TGCGGCAGAT CGGCAGGGTG 1201 CGGAAACTTC CCGAACAGCA GGGGCGGTATTTTACGGCAG ACAATTTGTC 1251 CAAAATACAG ATGTTCGCCC TGTCGAAGCT GCCCGACAAACGGGAGGCTT 1301 TGAGGGGGTT GGACAAGATT ATCGAAAAAC CGCCTGCCGG CAGTAATACA1351 GAGTTACAGG CAGAGGCATT GGTACAGCGG TCAGTTGTTT ACGATCGGCT 1401TGGCAAGCGG AAAAAAATGA TTTCAGATCT TGAAAGGGCG TTCAGGCTTG 1451 CACCCGATAACGCTCAGATT ATGAATAATC TGGGCTACAG CCTGCTTTCC 1501 GATTCCAAAC GTTTGGACGAAGGCTTCGCC CTGCTTCAGA CGGCATACCA 1551 AATCAACCCG GACGATACCG CTGTCAACGACAGCATAGGC TGGGCGTATT 1601 ACCTGAAANG CGACGCGGAA AGCGCGCTGC CGTATCTGCGGTATTCGTTT 1651 GAAAACGACC CCGAGCCCGA AGTTGCCGCC CATTTGGGCG AAGTGTTGTG1701 GGCATTGGGC GAACGCGATC AGGCGGTTGA CGTATGGACG CAGGCGGCAC 1751ACCTTACGGG AGACAAGAAA ATATGGCGGG AAACGCTCAA ACGTCACGGC 1801 ATCGCATTGCCCCAACCTTC CCGAAAACCT CGGAAATAA

This encodes a protein having amino acid sequence <SEQ ID 44>:

1 MLPARFTILS VLAAALLAGQ AYAAGAADAK PPKEVGKVFR KQQRYSEEEI 51 KNERARLAAVGERVNQIFTL LGXETALQKG QAGTALATYM LMLERTKSPE 101 VAERALEMAV SLNAFEQAEMIYQKWRQIEP IPGKAQKRAG WLRNVLRERG 151 NQHLDGLEEX LAQADEXQNR RVFLLLAQAAVQQDGLAQKA SKAVRRAALR 201 YEHLPEAAVA DVVFSVQXRE KEKAIGALQR LAKLDTEILPPTLMTLRLTA 251 RKYPEILDGF FEQTDTQNLS AVWQEMEIMN LVSLHRLDDA YARLNVLLER301 NPNADLYIQA AILAANRKEX ASVIDGYAEK AYGRGTGEQR GRAAMTAAMI 351YADRRDYTKV RQWLKKVSAP EYLFDKGVLA AAAAVELDXG RAALRQIGRV 401 RKLPEQQGRYFTADNLSKIQ MFALSKLPDK REALRGLDKI IEKPPAGSNT 451 ELQAEALVQR SVVYDRLGKRKKMISDLERA FRLAPDNAQI MNNLGYSLLS 501 DSKRLDEGFA LLQTAYQINP DDTAVNDSIGWAYYLKXDAE SALPYLRYSF 551 ENDPEPEVAA HLGEVLWALG ERDQAVDVWT QAAHLTGDKKIWRETLKRHG 601 IALPQPSRKP RK*

ORF9a and ORF9-1 show 95.3% identity in 614 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF9 shows 82.8% identity over a 163aa overlap with a predicted ORF(ORF9.ng) from N. gonorrhoeae:

The ORF9ng nucleotide sequence <SEQ ID 45> was predicted to encode aprotein having including acid sequence <SEQ ID 46>:

1 MIMLPARFTI LSVLAAALLA GQAYAAGAAD VELPKEVGKV LRKHRRYSEE 51EIKNERARLA AVGERVNRVF TLLGGETALQ KGQAGTALAT YMLMLERTKS 101PEVAERALEM AVSLNAFEQA EMIYQKWRQI EPIPGEAQKP AGWLRNVLKE 151GGNPHLDRLE EVPAQSDYVH QPMIFLLLVQ AAVQHGGVAQ KPSKAVRPAA 201YNYEVLPETA GADAVFCVQG PQYEKAIQSF PPCGRNPQTE NIAPPFNELF 251RPTARPISPK LLQRFFRTEP NLAKPFRPPG PEMETYQTGF PRPLTRNNPT

Amino acids 1-28 are a putative leader sequence, and 173-189 arepredicted to be a transmembrane domain.

Further sequence analysis revealed the complete length ORF9ng DNAsequence <SEQ ID 47>:

1 ATGTTACCCG CCCGTTTCAC TATTTTATCT GTCCTCGCAG CAGCCCTGCT 51TGCCGGACAG GCGTATGCTG CCGGCGCGGC GGATGTGGAG CTGCCGAAGG 101AAGTCGGAAA GGTTTTAAGG AAACATCGGC GTTACAGCGA GGAAGAAATC 151AAAAACGAAC GCGCACGGCT TGCGGCAGTG GGCGAACGGG TCAACAGGGT 201GTTTACGCTG TTGGGCGGTG AAACGGCTTT GCAGAAAGGG CAGGCGGGAA 251CGGCTCTGGC AACCTATATG CTGATGTTGG AACGCACAAA ATCCCCCGAA 301GTCGCCGAAC GCGCCTTGGA AATGGCCGTG TCGCTGAACG CGTTTGAACA 351GGCGGAAATG ATTTATCAGA AATGgcggca gatcgagcct ataCcgggtg 401aggcgcaaaa accgGcgggG tggctgcgga acgtattgaa ggaagggGGa 451aaTCAGCATC TGGAcgggtt gaaagaggTG CtggcgcaAT cggacgatGT 501GCAAAAAcgc aggaTATTTT TGCTGCTGGT GCAAGCCGCC GTGCagcagg 551gTGGGGTGGC TCAAAAAGCA TCGAAAGCGG TTCGCcgtgc GGcgttgaAG 601TATGAACATC TGCCcgaagc ggcggTTGCC GATGcggTGT TCGGCGTACA 651GGGACGCGAA AAGGAAAagg caaTCGAAGC TTTGCAGCGT TTGGCGAAGC 701TCGATACGGA AATATTGCCC CCCACTTTAA TGACGTTGCG TCTGACTGCA 751CGCAAATATC CCGAAATACT CGACGGCTTT TTCGAGCAGA CAGACACCCA 801AAACCTTTCG GCCGTCTGGC AGGAAATGGA AATTATGAAT CTGGTTTCCC 851TGCGTAAGCC GGATGATGCC TATGCGCGTT TGAACGTGCT GTTGGAACAC 901AACCCGAATG CAAACCTGTA TATTCAGGCG GCGATATTGG CGGCAAACCG 951AAAAGAAGGT GCGTCCGTTA TCGACGGCTA CGCCGAAAAG GCATACGGCA 1001GGGGGACGGG GGAACAGCGG GGCagggcgg cAATgacggc GGCGATGATA 1051TATGCCGACC GCAGGGATTA CGCCAAAGTC AGGCAGTGGT TGAAAAAAGT 1101GTCCGCGCCG GAATACCTGT TCGACAAAGG CGTGCTGGCG GCTGCGGCGG 1151CTGCCGAATT GGACGGAGGC CGGGCGGCTT TGCGGCAGAT CGGCAGGGTG 1201CGGAAACTTC CCGAACAGCA GGGGCGGTAT TTTACGGCAG ACAATTTGTC 1251CAAAATACAG ATGCTCGCCC TGTCGAAGCT GCCCGACAAA CGGGAAGCCC 1301TGATCGGGCT GAACAACATC ATCGCCAAAC TTTCGGCGGC GGGAAGCACG 1351GAACCTTTGG CGGAAGCATT GGCACAGCGT TCCATTATTT ACGaacAGTT 1401cggCAAACGG GGAAAAATGA TTGCCGACCT tgaAACcgcg CTCAAACTTA 1451CGCCCGATAA TGCACAAATT ATGAATAATC TGGGCTACAG CCTGCTTTCC 1501GATTCCAAAC GTTTGGACGA GGGTTTCGCC CTGCTTCAGA CGGCATACCA 1551AATCAACCCG GACGATACCG CCGTTAACGA CAGCATAGGC TGGGCGTATT 1601ACCTGAAAGG CGACgcggaA AGCGCGCTGC CGTATCTGcg gtattcgttt 1651gAAAACGACC CCGAGCCCGA AGTTGCCGCC CATTTGGGCG AAGTGTTGTG 1701GGCATTGGGC GAACGCGATC AGGCGGTTGA CGTATGGACG CAGGCGGCAC 1751ACCTTAGGGG AGACAAGAAA ATATGGCGGG AGACGCTCAA ACGCTACGGA 1801ATCGCCTTGC CCGAGCCTTC CCGAAAACCC CGGAAATAA

This encodes a protein having amino acid sequence <SEQ ID 48>:

1 MLPARFTILS VLAAALLAGQ AYAAGAADVE LPKEVGKVLR KHRRYSEEEI 51KNERARLAAV GERVNRVFTL LGGETALQKG QAGTALATYM LMLERTKSPE 101VAERALEMAV SLNAFEQAEM IYQKWRQIEP IPGEAQKPAG WLRNVLKEGG 151NQHLDGLKEV LAQSDDVQKR RIFLLLVQAA VQQGGVAQKA SKAVRRAALK 201YEHLPEAAVA DAVFGVQGRE KEKAIEALQR LAKLDTEILP PTLMTLRLTA 251RKYPEILDGF FEQTDTQNLS AVWQEMEIMN LVSLRKPDDA YARLNVLLEH 301NPNANLYIQA AILAANRKEG ASVIDGYAEK AYGRGTGEQR GRAAMTAAMI 351YADRRDYAKV RQWLKKVSAP EYLFDKGVLA AAAAAELDGG RAALRQIGRV 401RKLPEQQGRY FTADNLSKIQ MLALSKLPDK REALIGLNNI IAKLSAAGST 451EPLAEALAQR SIIYEQFGKR GKMIADLETA LKLTPDNAQI MNNLGYSLLS 501DSKRLDEGFA LLQTAYQINP DDTAVNDSIG WAYYLKGDAE SALPYLRYSF 551ENDPEPEVAA HLGEVLWALG ERDQAVDVWT QAAHLRGDKK IWRETLKRYG 601IALPEPSRKP RK*

ORF9ng and ORF9-1 show 88.1% identity in 614 aa overlap:

In addition, ORF9ng shows significant homology with a hypotheticalprotein from P. aeruginosa:

sp|P42810|YHE3_PSEAE HYPOTHETICAL 64.8 KD PROTEIN IN HEMM-HEMA INTERGENICREGION (ORF3)>gi|1072999|pir||S49376 hypothetical protein 3 - Pseudomonas aeruginosa>gi|557259 (X82071) orf3 [Pseudomonas aeruginosa] Length = 576 Score =128 bits (318), Expect = 1e−28 Identities = 138/587 (23%), Positives =228/587 (38%), Gaps = 125/587 (21%) Query: 67VFTLLGGETALQKGQAGTALATYMLMLERTKSPEVAERALEMAVSLNAFEQAEMIYQKWR 126+++LL  E A Q+ +   AL+ Y++  ++T+ P V+ERA  +A  L A ++A      W Sbjct: 53LYSLLVAELAGQRNRFDIALSNYVVQAQKTRDPGVSERAFRIAEYLGADQEALDTSLLWA 112 Query:127 QIEPIPGEAQKPAG--------------WLRNVLKEGGNQHLDGLKEVLAQSDDVQKRRI 172 + P   +AQ+ A               ++  VL   G+ H D L    A++D   +  + Sbjct: 113RSAPDNLDAQRAAAIQLARAGRYEESMVYMEKVLNGQGDTHFDFLALSAAETDPDTRAGL 172 Query:173 FXXXXXXXXXXXXXXXKASKAVRRAALKYEHLPEAAVADAVFGVQGREKEKAIEALQRLA 232                   ++      KY +  +     A+   Q    ++A+  L+  + Sbjct: 173L------------------QSFDHLLKKYPNNGQLLFGKALLLQQDGRPDEALTLLEDNS 214 Query:233 KLDTEILPPTLMTLRLTARK-----YPEILDGFFEQTDTQNLSAVWQEMEIMNLVSLRKP 287    E+ P  L +  L + K      P +  G  E  D + +   +  +    LV   + Sbjct: 215ASRHEVAPLLLRSRLLQSMKRSDEALPLLKAGIKEHPDDKRVRLAYARL----LVEQNRL 270 Query:288 DDAYARLNVLLEHNPN---------------------ANLYIQAAI-------------- 312DDA A    L++  P+                     A +Y++  + Sbjct: 271DDAKAEFAGLVQQFPDDDDDLRFSLALVCLEAQAWDEARIYLEELVERDSHVDAAHFNLG 330 Query:313 -LAANRKEGASVIDGYAEKAYGRGTGEQRGRAAMTAAMIYADRRDYAKVRQWLKKVSAPE 371 LA  +K+ A  +D YA+   G G      +   T  ++ A R D A  R    +   P+ Sbjct: 331RLAEEQKDTARALDEYAQ--VGPGNDFLPAQLRQTDVLLKAGRVDEAAQRLDKARSEQPD 388 Query:372 YLFDKXXXXXXXXXXXXXXXXXXRQIGRVRKLPEQQGRYFTADNLSKIQMLALSKLPDKR 431Y                                        A  L  I+  ALS    + Sbjct: 389Y----------------------------------------AIQLYLIEAEALSNNDQQE 408 Query:432 EALIGLNNIIAKLSAAGSTEPLAEALAQRSIIYEQFGKRGKMIADLETALKLTPDNAQIM 491+A   +   + +       E L   L  RS++ E+     +M  DL   +   PDNA  + Sbjct: 409KAWQAIQEGLKQYP-----EDL-NLLYTRSMLAEKRNDLAQMEKDLRFVIAREPDNAMAL 462 Query:492 NNLGYSLLSDSKRLDEGFALLQTAYQINPDDTAVNDSIGWAYYLKGDAESALPYLRYSFE 551N LGY+L   + R  E   L+  A+++NPDD A+ DS+GW  Y +G    A  YLR + + Sbjct: 463NALGYTLADRTTRYGEARELILKAHKLNPDDPAILDSMGWINYRQGKLADAERYLRQALQ 522 Query:552 NDPEPEVAAHLGEVLWALGERDQAVDVWTQAAHLRGDKKIWRETLKR 598   P+EVAAHLGEVLWA G +  A  +W +    + D  + R T+KR Sbjct: 523RYPDHEVAAHLGEVLWAQGRQGDARAIWREYLDKQPDSDVLRRTIKR 569gi|2983399 (AE000710) hypothetical protein [Aquifex aeolicus] Length =545 Score = 81.5 bits (198), Expect = 1e−14 Identities =61/198 (30%), Positives = 98/198 (48%), Gaps = 19/198 (9%) Query: 408GRYFTADNL-SKIQMLALSKLPDKREALIGLNNIIAKLSAAGSTEPLAEALAQ------- 459G Y  A  L  K ++LA    PDK+E L    +  +K         + + L + Sbjct: 335GNYEDAKRLIEKAKVLA----PDKKEILFLEADYYSKTKQYDKALEILKKLEKDYPNDSR 390 Query:460 ----RSIIYEQFGKRGKMIADLETALKLTPDNAQIMNNLGYSLLS--DSKRLDEGFALLQ 513     +I+Y+  G        L  A++L P+N    N LGYSLL     +R++E   L++ Sbjct: 391VYFMEAIVYDNLGDIKNAEKALRKAIELDPENPDYYNYLGYSLLLWYGKERVEEAEELIK 450 Query:514 TAYQINPDDTAVNDSIGWAYYLKGDAESALPYLRYSF-ENDPEPEVAAHLGEVLWALGER 572 A + +P++ A  DS+GW YYLKGD E A+ YL  +  E   +P V  H+G+VL  +G + Sbjct: 451KALEKDPENPAYIDSMGWVYYLKGDYERAMQYLLKALREAYDDPVVNEHVGDVLLKMGYK 510 Query:573 DQAVDVWTQAAHLRGDKK 590 ++A + + +A  L  + K Sbjct: 511EEARNYYERALKLLEEGK 528

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 7

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 49>:

1 AACCTCTACG CCGGCCCGCA GACCACATCC GTCATCGCAA ACATCGCCGA 51CAACCTGCAA CTGGCCAAAG ACTACGGCAA AGTACACTGG TTCGCCTCCC 101CGCTCTTCTG GCTCCTGAAC CAACTGCACA ACATCATCGG CAACTGGGGC 151TGGGCGATTA TCGTTTTAAC CATCATCGTC AAAGCCGTAC TGTATCCATT 201GACCAACGCC TCTTACCGCT CTATGGCGAA AATGCGTGCC GCCGCACCCA 251AACTGCAAGC CATCAAAGAG AAATACGGCG ACGACCGTAT GGCGCAACAA 301CAGGCGATGA TGCAGCTTTA CACAGACGAG AAAATCAACC CGaCTGGGCG 351GCTGCCTGCC TATGCTGTTG CAAATCCCCG TCTTCATCGG ATTGTATTGG 401GCATTGTTCG CCTCCGTAGA ATTGCGCCAG GCACCTTGGC TGGGTTGGAT 451TACCGACCTC AGCCGCGCCG ACCCCTACTA CATCCTGCCC ATCATTATGG 501CGGCAACGAT GTTCGCCCAA ACTTATCTGA ACCCGCCGCC GAcCGACCCG 551ATGCagGCGA AAATGATGAA AATCATGCCG TTGGTTTTCT CsGwCrTGTT 601CTTCTTCTTC CCTGCCGGks TGGTATTGTA CTGGGTAGTC AACAACCTCC 651TGACCATCGC CCAGCAATGG CACATCAACC GCAGCATCGA AAAACAACGC 701GCCCAAGGCG AAGTCGTTTC CTAA

This corresponds to the amino acid sequence <SEQ ID 50; ORF11>:

1 ..NLYAGPQTTS VIANIADNLQ LAKDYGKVHW FASPLFWLLN QLHNIIGNWG 51  WAIIVLTIIV KAVLYPLTNA SYRSMAKMRA AAPKLQAIKE KYGDDRMAQQ 101  QAMMQLYTDE KINPLGGCLP MLLQIPVFIG LYWALFASVE LRQAPWLGWI 151  TDLSRADPYY ILPIIMAATM FAQTYLNPPP TDPMQAKMMK IMPLVFSXXF 201  FFFPAGXVLY WVVNNLLTIA QQWHINRSIE KQRAQGEVVS *

Further sequence analysis revealed the complete DNA sequence <SEQ ID51>:

1 ATGGATTTTA AAAGACTCAC GGCGTTTTTC GCCATCGCGC TGGTGATTAT 51GATCGGCTGG GAAAAGATGT TCCCCACTCC GAAGCCAGTC CCCGCGCCCC 101AACAGGCAGC ACAACAACAG GCCGTAACCG CTTCCGCCGA AGCCGCGCTC 151GCGCCCGCAA CGCCGATTAC CGTAACGACC GACACGGTTC AAGCCGTCAT 201TGATGAAAAA AGCGGCGACC TGCGCCGGCT GACCCTGCTC AAATACAAAG 251CAACCGGCGA CGAAAATAAA CCGTTCATCC TGTTTGGCGA CGGCAAAGAA 301TACACCTACG TCGCCCAATC CGAACTTTTG GACGCGCAGG GCAACAACAT 351TCTAAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC AGCTTGGAAG 401GCGACAAAGT TGAAGTCCGC CTGAGCGCGC CTGAAACACG CGGTCTGAAA 451ATCGACAAAG TTTATACTTT CACCAAAGGC AGCTATCTGG TCAACGTCCG 501CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG AGCGCGGACT 551ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG TTACTTTACC 601CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA ACTTCCAAAA 651AGTCAGCTTT TCCGACTTGG ACGACGATGC CAAATCCGGC AAATCCGAGG 701CCGAATACAT CCGCAAAACC CCGACCGGCT GGCTCGGCAT GATTGAACAC 751CACTTCATGT CCACCTGGAT TCTCCAACCT AAAGGCAGAC AAAGCGTTTG 801CGCCGCAGGC GAGTGCAACA TCGACATCAA ACGCCGCAAC GACAAGCTGT 851ACAGCACCAG CGTCAGCGTG CCTTTAGCCG CCATCCAAAA CGGCGCGAAA 901GCCGAAGCCT CCATCAACCT CTACGCCGGC CCGCAGACCA CATCCGTCAT 951CGCAAACATC GCCGACAACC TGCAACTGGC CAAAGACTAC GGCAAAGTAC 1001ACTGGTTCGC CTCCCCGCTC TTCTGGCTCC TGAACCAACT GCACAACATC 1051ATCGGCAACT GGGGCTGGGC GATTATCGTT TTAACCATCA TCGTCAAAGC 1101CGTACTGTAT CCATTGACCA ACGCCTCTTA CCGCTCTATG GCGAAAATGC 1151GTGCCGCCGC ACCCAAACTG CAAGCCATCA AAGAGAAATA CGGCGACGAC 1201CGTATGGCGC AACAACAGGC GATGATGCAG CTTTACACAG ACGAGAAAAT 1251CAACCCGCTG GGCGGCTGCC TGCCTATGCT GTTGCAAATC CCCGTCTTCA 1301TCGGATTGTA TTGGGCATTG TTCGCCTCCG TAGAATTGCG CCAGGCACCT 1351TGGCTGGGTT GGATTACCGA CCTCAGCCGC GCCGACCCCT ACTACATCCT 1401GCCCATCATT ATGGCGGCAA CGATGTTCGC CCAAACTTAT CTGAACCCGC 1451CGCCGACCGA CCCGATGCAG GCGAAAATGA TGAAAATCAT GCCGTTGGTT 1501TTCTCCGTCA TGTTCTTCTT CTTCCCTGCC GGTCTGGTAT TGTACTGGGT 1551AGTCAACAAC CTCCTGACCA TCGCCCAGCA ATGGCACATC AACCGCAGCA 1601TCGAAAAACA ACGCGCCCAA GGCGAAGTCG TTTCCTAA

This corresponds to the amino acid sequence <SEQ ID 52; ORF11-1>:

1 MDFKRLTAFF AIALVIMIGW EKMFPTPKPV PAPQQAAQQQ AVTASAEAAL 51APATPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDENK PFILFGDGKE 101YTYVAQSELL DAQGNNILKG IGFSAPKKQY SLEGDKVEVR LSAPETRGLK 151IDKVYTFTKG SYLVNVRFDI ANGSGQTANL SADYRIVRDH SEPEGQGYFT 201HSYVGPVVYT PEGNFQKVSF SDLDDDAKSG KSEAEYIRKT PTGWLGMIEH 251HFMSTWILQP KGRQSVCAAG ECNIDIKRRN DKLYSTSVSV PLAAIQNGAK 301AEASINLYAG PQTTSVIANI ADNLQLAKDY GKVHWFASPL FWLLNQLHNI 351IGNWGWAIIV LTIIVKAVLY PLTNASYRSM AKMRAAAPKL QAIKEKYGDD 401RMAQQQAMMQ LYTDEKINPL GGCLPMLLQI PVFIGLYWAL FASVELRQAP 451WLGWITDLSR ADPYYILPII MAATMFAQTY LNPPPTDPMQ AKMMKIMPLV 501FSVMFFFFPA GLVLYWVVNN LLTIAQQWHI NRSIEKQRAQ GEVVS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a 60 kDa Inner-Membrane Protein (Accession P25754) ofPseudomonas putida

ORF11 and the 60 kDa protein show 58% aa identity in 229 aa overlap(BLASTp).

ORF11 2 LYAGPQTTSVIANIADNLQLAKDYGKVHWFASPLFWLLNQLHNIIGNWGWAIIVLTIIVK 61LYAGP+  S +  ++  L+L  DYG + + A P+FWLL  +H+++GNWGW+IIVLT+++K 60K 324LYAGPKIQSKLKELSPGLELTVDYGFLWFIAQPIFWLLQHIHSLLGNWGWSIIVLTMLIK 383 ORF1162 AVLYPLTNASYRSMAKMRAAAPKLQAIKEKYGDDRXXXXXXXXXLYTDEKINPLGGCLPM 121  ++PL+ ASYRSMA+MRA APKL A+KE++GDDR         LY  EKINPLGGCLP+ 60K 384GLFFPLSAASYRSMARMRAVAPKLAALKERFGDDRQKMSQAMMELYKKEKINPLGGCLPI 443 ORF11122 LLQIPVFIGLYWALFASVELRQAPWLGWITDLSRADPYYILPIIMAATMFAQTYLNPPPT 181L+Q+PVF+ LYW L  SVE+RQAPW+ WITDLS  DP++ILPIIM ATMF Q  LNP P 60K 444LVQMPVFLALYWVLLESVEMRQAPWILWITDLSIKDPFFILPIIMGATMFIQQRLNPTPP 503 ORF11182 DPMQAKMMKIMPLVXXXXXXXXPAGXVLYWVVNNLLTIAQQWHINRSIE 230 DPMQAK+MK+MP++       PAG VLYWVVNN L+I+QQW+I R IE 60K 504DPMQAKVMKMMPIIFTFFFLWFPAGLVLYWVVNNCLSISQQWYITRRIE 552Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF 11 shows 97.9% identity over a 240aa overlap with an ORF (ORF11a)from strain A of N. meningitidis:

The complete length ORF11a nucleotide sequence <SEQ ID 53> is:

ANGGATTTTA AAAGACTCAC NGNGTTTTTC GCCATCGCAC TGGTGATTAT 51GATCGGATNG NAAANGATGT TCCCCACTCC GAAGCCCGTC CCCGCGCCCC 101AACAGACGGC ACAACAACAG GCCGTAANCG CTTCCGCCGA AGCCGCGCTC 151GCGCCCGNAN CGCCGATTAC CGTAACGACC GACACGGTTC AAGCCGTCAT 201TGATGAAAAA AGCGGCGACC TGCGCCGGCT GACCCTGCTC AAATACAAAG 251CAACCGGCGA CNAAAATAAA CCGTTCATCC TGTTTGGCGA CGGCAAANAA 301TACACCTACN TCGCCCANTC CGAACTTTTG GACGCGCAGG GCAACAACAT 351TCTAAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC AGCTTGGAAG 401GCGACAAAGT TGAAGTCCGC CTGAGCGCAC CTGAAACACG CGGTCTGAAA 451ATCGACAAAG TTTATACTTT CACCAAAGGC AGCTATCTGG TCAACGTCCG 501CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG AGCGCGGACT 551ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG CTACTTTACC 601CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA ACTTCCAAAA 651AGTCAGCTTC TCCGACTTGG ACGACGATGC CAANTCCGGN AAATCCGAGG 701CCGAATACAT CCGCAAAACC CNGACCGGCT GGCTCGGCAT GATTGAACAC 751CACTTCATGT CCACCTGGAT CCTCCAACCC AAAGGCGGAC AAAGCGTTTG 801CGCCGCTGGC GACTGCNGTA TNGACATCAA ACGCCGCAAC GACAAGCTGT 851ACAGCACCAG CGTCAGCGTG CCTTTAGCCG CTATCCAAAA CGGTGCGAAA 901TCCNAAGCCT CCATCAACCT CTACGCCGGC CCACAGACCA CATCNGTTAT 951CGCAAACATC GCCGACAACC TGCAACTGGN CAAAGACTAC GGCAAAGTAC 1001ACTGGTTCGC CTCCCCCCTC TTTTGGCTTT TGAACCAACT GCACAACATC 1051ATCGGCAACT GGGGCTGGGC GATTATCGTT TTAACCATCA TCGTCAAAGC 1101CGTACTGTAT CCATTGACCA ACGCCTCTTA CCGTTCGATG GCGAAAATGC 1151GTGCCGCCGC GCCCAAACTG CAAGCCATCA AAGAGAAATA CGGCGACGAC 1201CGTATGGCGC AGCAACAAGC CATGATGCAG CTTTACACAG ACGAGAAAAT 1251CAACCCGCTG GGCGGCTGCC TGCCTATGCT GTTGCAAATC CCCGTCTTCA 1301TCGGATTGTA TTGGGCATTG TTCGCCTCCG TAGAATTGCG CCAGGCACCT 1351TGGCTGGGTT GGATTACCGA CCTCAGCCGC GCCGACCCNT ACTACATCCT 1401GCCCATCATT ATGGCGGCAA CGATGTTCGC CCAAACCTAT CTGAACCCGC 1451CGCCGACCGA CCCGATGCAG GCGAAAATGA TGAAAATCAT GCCTTTGGTT 1501NTNTCNNNNA NGTTCTTCNN CTTCCCTGCC GGTCTGGTAT TGTACTGGGT 1551GATCAACAAC CTCCTGACCA TCGCCCAGCA ATGGCACATC AACCGCAGCA 1601TCGAAAAACA ACGCGCCCAA GGCGAAGTCG TTTCCTAA

This encodes a protein having amino acid sequence <SEQ ID 54>:

1 XDFKRLTXFF AIALVIMIGX XXMFPTPKPV PAPQQTAQQQ AVXASAEAAL 51APXXPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDXNK PFILFGDGKX 101YTYXAXSELL DAQGNNILKG IGFSAPKKQY SLEGDKVEVR LSAPETRGLK 151IDKVYTFTKG SYLVNVRFDI ANGSGQTANL SADYRIVRDH SEPEGQGYFT 201HSYVGPVVYT PEGNFQKVSF SDLDDDAXSG KSEAEYIRKT XTGWLGMIEH 251HFMSTWILQP KGGQSVCAAG DCXXDIKRRN DKLYSTSVSV PLAAIQNGAK 301SXASINLYAG PQTTSVIANI ADNLQLXKDY GKVHWFASPL FWLLNQLHNI 351IGNWGWAIIV LTIIVKAVLY PLTNASYRSM AKMRAAAPKL QAIKEKYGDD 401RMAQQQAMMQ LYTDEKINPL GGCLPMLLQI PVFIGLYWAL FASVELRQAP 451WLGWITDLSR ADPYYILPII MAATMFAQTY LNPPPTDPMQ AKMMKIMPLV 501XSXXFFXFPA GLVLYWVINN LLTIAQQWHI NRSIEKQRAQ GEVVS*

ORF11a and ORF11-1 show 95.2% identity in 544 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF11 shows 96.3% identity over a 240aa overlap with a predicted ORF(ORF11.ng) from N. gonorrhoeae:

An ORF11ng nucleotide sequence <SEQ ID 55> was predicted to encode aprotein having amino acid sequence <SEQ ID 56>:

1 MAVNLYAGPQ TTSVIANIAD NLQLAKDYGK VHWFASPLFW LLNQLHNIIG 51NWGWAIVVLT IIVKAVLYPL TNASYRSMAK MRAAAPELQT IKEKYGDDRM 101AQQQAMMQLF EDEEINPLGG CLPMLLQIPV FIGLYWALFA SVELRQAPWL 151GWITDLSRAD PYYILPIIMA ATMFAQTYLN PPPTDPMQAK MMKIMPLVFS 201VMFFFFPAGL VLYWVVNNLL TIAQQWHINR SIEKQRAQGE VVS*

Further sequence analysis revealed the complete gonococcal DNA sequence<SEQ ID 57> to be:

1 ATGGATTTTA AAAGACTCAC GGCGTTTTTC GCCATCGCGC TGGTGATTAT 51GATCGGCTGG GAAAAAATGT TCCCCACCCC GAAACCCGTC CCCGCGCCCC 101AACAGGCGGC ACAAAAACAG GCAGCAACCG CTTCCGCCGA AGCCGCGCTC 151GCGCCCGCAA CGCCGATTAC CGTAACGACC GACACGGTTC AAGCCGTTAT 201TGATGAAAAA AGTGGCGACC TGCGCCGGCT GACCCTGCTC AAATACAAAG 251CAACCGGCGA CGAAAACAAA CCGTTCGTCC TGTTTGGCGA CGGCAAAGAA 301TACACCTACG TCGCCCAATC CGAACTTTTG GACGCGCAGG GCAACAACAT 351TCTGAAAGGC ATCGGCTTTA GCGCACCGAA AAAACAGTAC ACCCTCAACG 401GCGACACAGT CGAAGTCCGC CTGAGCGCGC CCGAAACCAA CGGACTGAAA 451ATCGACAAAG TCTATACCTT TACCAAAGAC AGCTATCTGG TCAACGTCCG 501CTTCGACATC GCCAACGGCA GCGGTCAAAC CGCCAACCTG AGCGCGGACT 551ACCGCATCGT CCGCGACCAC AGCGAACCCG AGGGTCAAGG CTACTTTACC 601CACTCTTACG TCGGCCCTGT TGTTTATACC CCTGAAGGCA ACTTCCAAAA 651AGTCAGCTTC TCCgacTTgg acgACGATGC gaaaTccggc aaATccgagg 701ccgaatacaT CCGCAAAACC ccgaccggtt ggctcggcat gattgaacac 751cacttcatgt ccacctggat cctccAAcct aaaggcggcc aaaacgtttg 801cgcccaggga gactgccgta tcgacattaa aCgccgcaac gacaagctgt 851acagcgcaag cgtcagcgtg cctttaaccg ctatcccaac ccgggggcca 901aaaccgaaaa tggcggTCAA CCTGTATGCC GGTCCGCAAA CCACATCCGT 951TATCGCAAAC ATCGCcgacA ACCTGCAACT GGCAAAAGAC TACGGTAAAG 1001TACACTGGTT CGCATCGCCG CTCTTCTGGC TCCTGAACCA ACTGCACAAC 1051ATTATCGGCA ACTGGGGCTG GGCAATCGTC GTTTTGACCA TCATCGTCAA 1101AGCCGTACTG TATCCATTGA CCAACGcctc ctACCGTTCG ATGGCGAAAA 1151TGCGTGccgc cgcacCcaaA CTGCAGACCA TCAAAGAAAA ATAcgGCGAC 1201GACCGTATGG CGCAACAGCA AGCGATGATG CAGCTTTACA AAgacgAGAA 1251AATCAACCCG CTGGGCGGCT GTctgcctat gctgttgCAA ATCCCCGTCT 1301TCATCGGCTT GTACTGGGCA TTGTTCGCCT CCGTAGAATT GCGCCAGGCA 1351CCTTGGCTGG GCTGGATTAC CGACCTCAGC CGCGCCGACC CCTACTACAT 1401CCTGCCCATC ATTATGGCGG CAACGATGTT CGCCCAAACC TATCTGAACC 1451CGCCGCCGAC CGACCCGATG CAGGCGAAAA TGATGAAAAT CATGCCGTTG 1501GTTTTCTCCG TCATGTTCTT CTTCTTCCCT GCCGGTTTGG TTCTCTACTG 1551GGTGGTCAAC AACCTCCTGA CCATCGCCCA GCAGTGGCAC ATCAACCGCA 1601GCATCGAAAA ACAACGCGCC CAAGGCGAAG TCGTTTCCTA A

This encodes a protein having amino acid sequence <SEQ ID 58;ORF11ng-1>:

1 MDFKRLTAFF AIALVIMIGW EKMFPTPKPV PAPQQAAQKQ AATASAEAAL 51APATPITVTT DTVQAVIDEK SGDLRRLTLL KYKATGDENK PFVLFGDGKE 101YTYVAQSELL DAQGNNILKG IGFSAPKKQY TLNGDTVEVR LSAPETNGLK 151IDKVYTFTKD SYLVNVRFDI ANGSGQTANL SADYRIVRDH SEPEGQGYFT 201HSYVGPVVYT PEGNFQKVSF SDLDDDAKSG KSEAEYIRKT PTGWLGMIEH 251HFMSTWILQP KGGQNVCAQG DCRIDIKRRN DKLYSASVSV PLTAIPTRGP 301KPKMAVNLYA GPQTTSVIAN IADNLQLAKD YGKVHWFASP LFWLLNQLHN 351IIGNWGWAIV VLTIIVKAVL YPLTNASYRS MAKMRAAAPK LQTIKEKYGD 401DRMAQQQAMM QLYKDEKINP LGGCLPMLLQ IPVFIGLYWA LFASVELRQA 451PWLGWITDLS RADPYYILPI IMAATMFAQT YLNPPPTDPM QAKMMKIMPL 501VFSVMFFFFP AGLVLYWVVN NLLTIAQQWH INRSIEKQRA QGEVVS*

ORF11ng-1 and ORF11-1 shown 95.1% identity in 546 aa overlap:

In addition, ORF11ng-1 shows significant homology with an inner-membraneprotein from the database (accession number p25754):

Based on this analysis, including the homology to an inner-membraneprotein from P. putida and the predicted transmembrane domains (seen inboth the meningococcal and gonoccal proteins), it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 8

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 59>:

1 ..GCCGTCTTAA TCATCGAATT ATTGACGGGA ACGGTTTATC TTTTGGTTGT 51  NAGCGCGGCT TTGGCGGGTT CGGGCATTGC TTACGGGCTG ACCGGCAGTA 101  CGCCTGCCGC CGTCTTGACC GNCGCTCTGC TTTCCGCGCT GGGTATTTNG 151  TTCGTACACG CCAAAACCGC CGTTAGAAAA GTTGAAACGG ATTCATATCA 201  GGATTTGGAT GCCGGACAAT ATGTCGAAAT CCTCCGNCAC ACAGGCGGCA 251  ACCGTTACGA AGTT.TTTAT CGCGGTACG. ACTGGCAGGC TCAAAATACG 301  GGGCAAGAAG AGCTTGAACC AGGAACTCGC GCCCTCATTG TCCGCAAGGA 351  AGGCAACCTT CTTATTATCA CACACCCTTA A

This corresponds to the amino acid sequence <SEQ ID 60; ORF13>:

1 ..AVLIIELLTG TVYLLVVSAA LAGSGIAYGL TGSTPAAVLT XALLSALGIX 51  FVHAKTAVRK VETDSYQDLD AGQYVEILRH TGGNRYEVXY RGTXWQAQNT 101  GQEELEPGTR ALIVRKEGNL LIITHP*

Further sequence analysis elaborated the DNA sequence slightly <SEQ ID61>:

1 ..GCCGTCTTAA TCATCGAATT ATTGACGGGA ACGGTTTATC TTTTGGTTGT 51  nAGCGCGGCT TTGGCGGGTT CGGGCATTGC TTACGGGCTG ACCGGCAGTA 101  CGCCTGCCGC CGTCTTGACC GnCGCTCTGC TTTCCGCGCT GGGTATTTnG 151  TTCGTACACG CCAAAACCGC CGTTAGAAAA GTTGAAACGG ATTCATATCA 201  GGATTTGGAT GCCGGACAAT ATGTCGAAAT CCTCCGACAC ACAGGCGGCA 251  ACCGTTACGA AGTTTTtTAT CGCGGTACGc ACTGGCAGGC TCAAAATACG 301  GGGCAAGAAG AGCTTGAACC AGGAACTCGC GCCCTCATTG TCCGCAAGGA 351  AGGCAACCTT CTTATTATCA CACACCCTTA A

This corresponds to the amino acid sequence <SEQ ID 62; ORF13-1>:

1 ..AVLIIELLTG TVYLLVVSAA LAGSGIAYGL TGSTPAAVLT XALLSALGIX 51  FVHAKTAVRK VETDSYQDLD AGQYVEILRH TGGNRYEVFY RGTHWQAQNT 101  GQEELEPGTR ALIVRKEGNL LIITHP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF13 shows 92.9% identity over a 126aa overlap with an ORF (ORF13a)from strain A of N. meningitidis:

The complete length ORF13a nucleotide sequence <SEQ ID 63> is:

1 ATGACTGTAT GGTTTGTTGC CGCTGTTGCC GTCTTAATCA TCGAATTATT 51GACGGGAACG GTTTATCTTT TGGTTGTCAG CGCGGCTTTG GCGGGTTCGG 101GCATTGCTTA CGGGCTGACC GGCAGCACGC CTGCCGCCGT CTTGACCGCC 151GCTCTGCTTT CCGCGCTGGG TATTTGGTTC GTACACGCCA AAACCGCCGT 201GGGAAAAGTT GAAACGGATT CATATCAGGA TTTGGATGCC GGGCAATATG 251CCGAAATCCT CCGGCACGCA GGCGGCAACC GTTACGAAGT TTTTTATCGC 301GGTACGCACT GGCAGGCTCA AAATACGGGG CAAGAAGAGC TTGAACCAGG 351AACGCGCGCC CTAATCGTCC GCAAGGAAGG CAACCTTCTT ATCATCGCAA 401 AACCTTAA

This encodes a protein having amino acid sequence <SEQ ID 64>:

1 MTVWFVAAVA VLIIELLTGT VYLLVVSAAL AGSGIAYGLT GSTPAAVLTA 51ALLSALGIWF VHAKTAVGKV ETDSYQDLDA GQYAEILRHA GGNRYEVFYR 101GTHWQAQNTG QEELEPGTRA LIVRKEGNLL IIAKP*

ORF13a and ORF13-1 show 94.4% identity in 126 aa overlap

Homology with a Predicted ORF from N. gonorrhoeae

ORF13 shows 89.7% identity over a 126aa overlap with a predicted ORF(ORF13.ng) from N. gonorrhoeae:

The complete length ORF13ng nucleotide sequence <SEQ ID 65> is:

1 ATGACTGTAT GGTTTGTTGC CGCTGTTGCC GTCTTAATCA TCGAATTATT 51GACGGGAACG GTTTATCTTT TGGTTGTCAG CGCGGCTTTG GCGGGTTCGG 101GCATTGCCTA CGGGCTGACT GGCAGCACGC CTGCCGCCGT CTTGACCGCC 151GCACTGCTTT CCGCGCTGGG CATTTGGTTC GTACATGCCA AAACCGCCGT 201GGGAAAAGTT GAAACGGATT CATATCAGGA TTTGGATACC GGAAAATATG 251CCGAAATCCT CCGATACACA GGCGGCAACC GTTACGAAGT TTTTTATCGC 301GGTACGCACT GGCAGGCGCA AAATACGGGG CAGGAAGTGT TTGAACCGGG 351AACGCGCGCC CTCATCGTCC GCAAAGAAGG TAACCTTCTT ATCATCGCAA 401 ACCCTTAA

This encodes a protein having amino acid sequence <SEQ ID 66>:

1 MTVWFVAAVA VLIIELLTGT VYLLVVSAAL AGSGIAYGLT GSTPAAVLTA 51ALLSALGIWF VHAKTAVGKV ETDSYQDLDT GKYAEILRYT GGNRYEVFYR 101GTHWQAQNTG QEVFEPGTRA LIVRKEGNLL IIANP*

ORF13ng shows 91.3% identity in 126 aa overlap with ORF13-1:

Based on this analysis, including the extensive leader sequence in thisprotein, it is predicted that ORF13 and ORF13ng are likely to be outermembrane proteins. It is thus predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 9

The following DNA sequence was identified in N. meningitidis <SEQ ID67>:

1 ATGTwTGATT TCGGTTTrGG CGArCTGGTT TTTGTCGGCA TTATCGCCCT 51GATwGtCCTC GGCCCCGAAC GCsTGCCCGA GGCCGCCCGC AyCGCCGGAC 101GGcTCATCGG CAGGCTGCAA CGCTTTGTCG GcAGCGTCAA ACAGGAATTT 151GACACTCAAA TCGAACTGGA AGAACTGAGG AAGGCAAAGC AGGAATTTGA 201AGCTGCCGcC GCTCAGGTTC GAGACAGCCT CAAAGAAACC GGTACGGATA 251TGGAAGGCAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC TTGGGAAAAA 301CTGCCCGAAC AGCGGACACC TGCCGATTTC GGTGTCGATG AAAACGGCAA 351TCCGCT.TCC CGATGCGGCA AACACCCTAT CAGACGGCAT TTCCGACGTT 401 ATGCCGTC..

This corresponds to the amino acid sequence <SEQ ID 68; ORF2>:

1 MXDFGLGELV FVGIIALIVL GPERXPEAAR XAGRLIGRLQ RFVGSVKQEF 51 DTQIELEELRKAKQEFEAAA AQVRDSLKET GTDMEGNLHD ISDGLKPWEK 101 LPEQRTPADF GVDENGNPXSRCGKHPIRRH FRRYAV..

Further work revealed the complete nucleotide sequence <SEQ ID 69>:

1 ATGTTTGATT TCGGTTTGGG CGAGCTGGTT TTTGTCGGCA TTATCGCCCT 51 GATTGTCCTCGGCCCCGAAC GCCTGCCCGA GGCCGCCCGC ACCGCCGGAC 101 GGCTCATCGG CAGGCTGCAACGCTTTGTCG GCAGCGTCAA ACAGGAATTT 151 GACACTCAAA TCGAACTGGA AGAACTGAGGAAGGCAAAGC AGGAATTTGA 201 AGCTGCCGCC GCTCAGGTTC GAGACAGCCT CAAAGAAACCGGTACGGATA 251 TGGAAGGCAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC TTGGGAAAAA301 CTGCCCGAAC AGCGGACACC TGCCGATTTC GGTGTCGATG AAAACGGCAA 351TCCGCTTCCC GATGCGGCAA ACACCCTATC AGACGGCATT TCCGACGTTA 401 TGCCGTCCGAACGTTCCTAC GCTTCCGCCG AAACCCTTGG GGACAGCGGG 451 CAAACCGGCA GTACAGCCGAACCCGCGGAA ACCGACCAAG ACCGCGCATG 501 GCGGGAATAC CTGACTGCTT CTGCCGCCGCACCCGTCGTA CAGACCGTCG 551 AAGTCAGCTA TATCGATACT GCTGTTGAAA CGCCTGTTCCGCACACCACT 601 TCCCTGCGCA AACAGGCAAT AAGCCGCAAA CGCGATTTTC GTCCGAAACA651 CCGCGCCAAA CCTAAATTGC GCGTCCGTAA ATCATAA

This corresponds to the amino acid sequence <SEQ ID 70; ORF2-1>:

1 MFDFGLGELV FVGIIALIVL GPERLPEAAR TAGRLIGRLQ RFVGSVKQEF 51 DTQIELEELRKAKQEFEAAA AQVRDSLKET GTDMEGNLHD ISDGLKPWEK 101 LPEQRTPADF GVDENGNPLPDAANTLSDGI SDVMPSERSY ASAETLGDSG 151 QTGSTAEPAE TDQDRAWREY LTASAAAPVVQTVEVSYIDT AVETPVPHTT 201 SLRKQAISRK RDFRPKHRAK PKLRVRKS*

Further work identified the corresponding gene in strain A of N.meningitidis <SEQ ID 71>:

1 ATGTTTGATT TCGGTTTGGG CGAGCTGGTT TTTGTCGGCA TTATCGCCCT 51 GATTGTCCTCGGCCCCGAAC GCCTGCCCGA GGCCGCCCGC ACCGCCGGAC 101 GGCTCATCGG CAGGCTGCAACGCTTTGTCG GCAGCGTCAA ACAGGAATTT 151 GACACGCAAA TCGAACTGGA AGAACTAAGGAAGGCAAAGC AGGAATTTGA 201 AGCTGCCGCT GCTCAGGTTC GAGACAGCCT CAAAGAAACCGGTACGGATA 251 TGGAGGGTAA TCTGCACGAC ATTTCCGACG GTCTGAAGCC TTGGGAAAAA301 CTGCCCGAAC AGCGCACGCC TGCTGATTTC GGTGTCGATG AAAACGGCAA 351TCCCTTTCCC GATGCGGCAA ACACCCTATT AGACGGCATT TCCGACGTTA 401 TGCCGTCCGAACGTTCCTAC GCTTCCGCCG AAACCCTTGG GGACAGCGGG 451 CAAACCGGCA GTACAGCCGAACCCGCGGAA ACCGACCAAG ACCGTGCATG 501 GCGGGAATAC CTGACTGCTT CTGCCGCCGCACCCGTCGTA CAGACCGTCG 551 AAGTCAGCTA TATCGATACC GCTGTTGAAA CCCCTGTTCCGCATACCACT 601 TCGCTGCGTA AACAGGCAAT AAGCCGCAAA CGCGATTTGC GTCCTAAATC651 CCGCGCCAAA CCTAAATTGC GCGTCCGTAA ATCATAA

This encodes a protein having amino acid sequence <SEQ ID 72; ORF2a>:

1 MFDFGLGELV FVGIIALIVL GPERLPEAAR TAGRLIGRLQ RFVGSVKQEF 51 DTQIELEELRKAKQEFEAAA AQVRDSLKET GTDMEGNLHD ISDGLKPWEK 101 LPEQRTPADF GVDENGNPFPDAANTLLDGI SDVMPSERSY ASAETLGDSG 151 QTGSTAEPAE TDQDRAWREY LTASAAAPVVQTVEVSYIDT AVETPVPHTT 201 SLRKQAISRK RDLRPKSRAK PKLRVRKS*

The originally-identified partial strain B sequence (ORF2) shows 97.5%identity over a 118aa overlap with ORF2a:

The complete strain B sequence (ORF2-1) and ORF2a show 98.2% identity in228 aa overlap:

Further work identified a partial DNA sequence <SEQ ID 73> in N.gonorrhoeae encoding the following amino acid sequence <SEQ ID 74;ORF2ng>:

1 MFDFGLGELI FVGIIALIVL GPERLPEAAR TAGRLIGRLQ RFVGSVKQEL 51 DTQIELEELRKVKQAFEAAA AQVRDSLKET DTDMQNSLHD ISDGLKPWEK 101 LPEQRTPADF GVDEKGNSLSRYGKHRIRRH FRRYAV*

Further work identified the complete gonococcal gene sequence <SEQ ID75>:

1 ATGTTTGATT TCGGTTTGGG CGAGCTGATT TTTGTCGGCA TTATCGCCCT 51 GATTGTCCTTGGTCCAGAAC GCCTGCCCGA AGCCGCCCGC ACTGCCGGAC 101 GGCTTATCGG CAGGCTGCAACGCTTTGTAG GAAGCGTCAA ACAAGAACTT 151 GACACTCAAA TCGAACTGGA AGAGCTGAGGAAGGTCAAGC AGGCATTCGA 201 AGCTGCCGCC GCTCAGGTTC GAGACAGCCT CAAAGAAACCGATACGGATA 251 TGCAGAACAG TCTGCACGAC ATTTCCGACG GTCTGAAGCC TTGGGAAAAA301 CTGCCCGAAC AGCGCACGCc tgccgatttc gGTGTCGATg AAAacggcaa 351tccccttccc gATACGGCAA ACACCGTATC AGACGGCATT TCCGACGTTA 401 TGCCGTCTGAACGTTCCGAT ACTtccgcCG AAACCCTTGG GGACGACAGG 451 CAAACCGGCA GTACAGCCGAACCTGCGGAA ACCGACAAAG ACCGCGCATG 501 GCGGGAATAC CTGactgctt ctgccgccgcacctgtcgta Cagagggccg 551 tcgaagtcag ctaTATCGAT ACTGCTGTTG AAacgcctgTtccgcaCacc 601 acttccctgc gcaAACAGGC AATAAACCGC AAACGCGATT TttgtccgaA651 ACACCGCGCc aAACCGAAat tgcgcgtcCG TAAATCATAA

This encodes a protein having the amino acid sequence <SEQ ID 76;ORF2ng-1>:

1 MFDFGLGELI FVGIIALIVL GPERLPEAAR TAGRLIGRLQ RFVGSVKQEL 51 DTQIELEELRKVKQAFEAAA AQVRDSLKET DTDMQNSLHD ISDGLKPWEK 101 LPEQRTPADF GVDENGNPLPDTANTVSDGI SDVMPSERSD TSAETLGDDR 151 QTGSTAEPAE TDKDRAWREY LTASAAAPVVQRAVEVSYID TAVETPVPHT 201 TSLRKQAINR KRDFCPKHRA KPKLRVRKS*

The originally-identified partial strain B sequence (ORF2) shows 87.5%identity over a 136aa overlap with ORF2ng:

The complete strain B and gonococcal sequences (ORF2-1 & ORF2ng-1) show91.7% identity in 229 aa overlap:

Computer analysis of these amino acid sequences indicates atransmembrane region (underlined), and also revealed homology (59%identity) between the gonococcal sequence and the TatB protein of E.coli:

gnl|PID|e1292181 (AJ005830) TatB protein [Escherichia coli] Length = 171Score = 56.6 bits (134), Expect = 1e−07 Identities =30/88 (34%), Positives = 52/88 (59%), Gaps = 1/88 (1%)Query:  1 MFDFGLGELIFVGIIALIVLGPERLPEAARTAGRLIGRLQRFVGSVKQELDTQIELEELR 60          MFD G  EL+ V II L+VLGP+RLP A +T    I  L+     +V+ EL +++L+E  +Sbjct:  1 MFDIGFSELLLVFIIGLVVLGPQRLPVAVKTVAGWIRALRSLATTVQNELTQELKLQEFQ 60Query: 61 -KVKQAFEAAAAQVRDSLKETDTDMQNS 87              +K+  +A+   +  LK +  +++ + Sbjct: 61 DSLKKVEKASLTNLTPELKASMDELRQA 88

Based on this analysis, it was predicted that ORF2, ORF2a and ORF2ng arelikely to be membrane proteins and so the proteins from N. meningitidisand N. gonorrhoeae, and their epitopes, could be useful antigens forvaccines or diagnostics, or for raising antibodies.

ORF2-1 (16 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 3A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 3B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for Westernblots (FIG. 3C), ELISA (positive result), and FACS analysis (FIG. 3D).These experiments confirm that ORF37-1 is a surface-exposed protein, andthat it is a useful immunogen.

Example 10

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 77>:

1 ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT TTATTTTATC 51 CGC.TGCGGGACACTGACAG GTATTCCATC GCATGGCGgA GkTAAACgCT 101 TTgCGGTCGA ACAAGAACTTGTGGCCGCTT CTGCCAGAGC TGCCGTTAAA 151 GACATGGATT TACAGGCATT ACACGGACGAAAAGTTGCAT TGTACATTGC 201 CACTATGGGC GACCAAGGTT CAGGcAGTTT GACAGGGGGGTCGCTACTCC 251 ATTGATGCAC kGrTwCsTGG CGAATACATA AACAGCCCTG CCGTCCGTAC301 CGATTACACC TATCCACGTT ACGAAACCAC CGCTGAAACA ACATCAGGCG 351GTTTGACAGG TTTAACCACT TCTTTATCTA CACTTAATGC CCCTGCACTC 401 TCTCGCACCCAATCAGACGG TAGCGGAAGT AAAAGCAGTC TGGGCTTAAA 451 TATTGGCGGG ATGGGGGATTATCGAAATGA AACCTTGACG ACTAACCCGC 501 GCGACACTGC CTTTCTTTCC CACTTGGTACAGACCGTATT TTTCCTGCGC 551 GGCATAGACG TTGTTTCTCC TGCCAATGCC GATACAGATGTGTTTATTAA 601 CATCGACGTA TTCGGAACGA TACGCAACAG AACCGAAATG..

This corresponds to the amino acid sequence <SEQ ID 78; ORF15>:

1 MQARLLIPIL FSVFILSACG TLTGIPSHGG XKRFAVEQEL VAASARAAVK 51 DMDLQALHGRKVALYIATMG DQGSGSLTGG RYSIDAXXXG EYINSPAVRT 101 DYTYPRYETT AETTSGGLTGLTTSLSTLNA PALSRTQSDG SGSKSSLGLN 151 IGGMGDYRNE TLTTNPRDTA FLSHLVQTVFFLRGIDVVSP ANADTDVFIN 201 IDVFGTIRNR TEM..

Further work revealed the complete nucleotide sequence <SEQ ID 79>:

1 ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT TTATTTTATC 51 CGCCTGCGGGACACTGACAG GTATTCCATC GCATGGCGGA GGTAAACGCT 101 TTGCGGTCGA ACAAGAACTTGTGGCCGCTT CTGCCAGAGC TGCCGTTAAA 151 GACATGGATT TACAGGCATT ACACGGACGAAAAGTTGCAT TGTACATTGC 201 CACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGTCGCTACTCCA 251 TTGATGCACT GATTCGTGGC GAATACATAA ACAGCCCTGC CGTCCGTACC301 GATTACACCT ATCCACGTTA CGAAACCACC GCTGAAACAA CATCAGGCGG 351TTTGACAGGT TTAACCACTT CTTTATCTAC ACTTAATGCC CCTGCACTCT 401 CTCGCACCCAATCAGACGGT AGCGGAAGTA AAAGCAGTCT GGGCTTAAAT 451 ATTGGCGGGA TGGGGGATTATCGAAATGAA ACCTTGACGA CTAACCCGCG 501 CGACACTGCC TTTCTTTCCC ACTTGGTACAGACCGTATTT TTCCTGCGCG 551 GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACAGATGTGTTTATTAAC 601 ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC ACCTATACAA651 TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC GCAGTAGACA 701GAACCAATAA AAAATTGCTC ATCAAACCAA AAACCAATGC GTTTGAAGCT 751 GCCTATAAAGAAAATTACGC ATTGTGGATG GGGCCGTATA AAGTAAGCAA 801 AGGAATTAAA CCGACGGAAGGATTAATGGT CGATTTCTCC GATATCCGAC 851 CATACGGCAA TCATACGGGT AACTCCGCCCCATCCGTAGA GGCTGATAAC 901 AGTCATGAGG GGTATGGATA CAGCGATGAA GTAGTGCGACAACATAGACA 951 AGGACAACCT TGA

This corresponds to the amino acid sequence <SEQ ID 80; ORF15-1>:

1 MQARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL VAASARAAVK 51 DMDLQALHGRKVALYIATMG DQGSGSLTGG RYSIDALIRG EYINSPAVRT 101 DYTYPRYETT AETTSGGLTGLTTSLSTLNA PALSRTQSDG SGSKSSLGLN 151 IGGMGDYRNE TLTTNPRDTA FLSHLVQTVFFLRGIDVVSP ANADTDVFIN 201 IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLLIKPKTNAFEA 251 AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIRPYGNHTG NSAPSVEADN301 SHEGYGYSDE VVRQHRQGQP *

Further work identified the corresponding gene in strain A of N.meningitidis <SEQ ID 81>:

1 ATGCAAGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT TTATTTTATC 51 CGCCTGCGGGACACTGACAG GTATTCCATC GCATGGCGGA GGTAAACGCT 101 TTGCGGTCGA ACAAGAACTTGTGGCCGCTT CTGCCAGAGC TGCCGTTAAA 151 GACATGGATT TACAGGCATT ACACGGACGAAAAGTTGCAT TGTACATTGC 201 AACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGTCGCTACTCCA 251 TTGATGCACT GATTCGTGGC GAATACATAA ACAGCCCTGC CGTCCGTACC301 GATTACACCT ATCCACGTTA CGAAACCACC GCTGAAACAA CATCAGGCGG 351TTTGACAGGT TTAACCACTT CTTTATCTAC ACTTAATGCC CCTGCACTCT 401 CGCGCACCCAATCAGACGGT AGCGGAAGTA AAAGCAGTCT GGGCTTAAAT 451 ATTGGCGGGA TGGGGGATTATCGAAATGAA ACCTTGACGA CTAACCCGCG 501 CGACACTGCC TTTCTTTCCC ACTTGGTACAGACCGTATTT TTCCTGCGCG 551 GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACGGATGTGTTTATTAAC 601 ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC ACCTATACAA651 TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC GCAGTAGACA 701GAACCAATAA AAAATTGCTC ATCAAACCAA AAACCAATGC GTTTGAAGCT 751 GCCTATAAAGAAAATTACGC ATTGTGGATG GGACCGTATA AAGTAAGCAA 801 AGGAATTAAA CCGACAGAAGGATTAATGGT CGATTTCTCC GATATCCAAC 851 CATACGGCAA TCATATGGGT AACTCTGCCCCATCCGTAGA GGCTGATAAC 901 AGTCATGAGG GGTATGGATA CAGCGATGAA GCAGTGCGACGACATAGACA 951 AGGGCAACCT TGA

This encodes a protein having amino acid sequence <SEQ ID 82; ORF15a>:

1 MQARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL VAASARAAVK 51 DMDLQALHGRKVALYIATMG DQGSGSLTGG RYSIDALIRG EYINSPAVRT 101 DYTYPRYETT AETTSGGLTGLTTSLSTLNA PALSRTQSDG SGSKSSLGLN 151 IGGMGDYRNE TLTTNPRDTA FLSHLVQTVFFLRGIDVVSP ANADTDVFIN 201 IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLLIKPKTNAFEA 251 AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIQPYGNHMG NSAPSVEADN301 SHEGYGYSDE AVRRHRQGQP *

The originally-identified partial strain B sequence (ORF15) shows 98.1%identity over a 213aa overlap with ORF15a:

The complete strain B sequence (ORF15-1) and ORF15a show 98.8% identityin 320 aa overlap:

Further work identified the corresponding gene in N. gonorrhoeae <SEQ ID83>:

1 ATGCGGGCAC GGCTGCTGAT ACCTATTCTT TTTTCAGTTT TTATTTTATC 51 CGCCTGCGGGACACTGACAG GTATTCCATC GCATGGCGGA GGCAAACGCT 101 TCGCGGTCGA ACAAGAACTTGTGGCCGCTT CTGCCAGAGC TGCCGTTAAA 151 GACATGGATT TACAGGCATT ACACGGACGAAAAGTTGCAT TGTACATTGC 201 AACTATGGGC GACCAAGGTT CAGGCAGTTT GACAGGGGGTCGCTACTCCA 251 TTGATGCACT GATTCGCGGC GAATACATAA ACAGCCCTGC CGTCCGCACC301 GATTACACCT ATCCGCGTTA CGAAACCACC GCTGAAACAA CATCAGGCGG 351TTTGACGGGT TTAACCACTT CTTTATCTAC ACTTAATGCC CCTGCACTCT 401 CGCGCACCCAATCAGACGGT AGCGGAAGTA GGAGCAGTCT GGGCTTAAAT 451 ATTGGCGGGA TGGGGGATTATCGAAATGAA ACCTTGACGA CCAACCCGCG 501 CGACACTGCC TTTCTTTCCC ACTTGGTGCAGACCGTATTT TTCCTGCGCG 551 GCATAGACGT TGTTTCTCCT GCCAATGCCG ATACAGATGTGTTTATTAAC 601 ATCGACGTAT TCGGAACGAT ACGCAACAGA ACCGAAATGC ACCTATACAA651 TGCCGAAACA CTGAAAGCCC AAACAAAACT GGAATATTTC GCAGTAGACA 701GAACCAATAA AAAATTGCTC ATCAAACCCA AAACCAATGC GTTTGAAGCT 751 GCCTATAAAGAAAATTACGC ATTGTGGATG GGGCCGTATA AAGTAAGCAA 801 AGGAATCAAA CCGACGGAAGGATTGATGGT CGATTTCTCC GATATCCAAC 851 CATACGGCAA TCATACGGGT AACTCCGCCCCATCCGTAGA GGCTGATAAC 901 AGTCATGAGG GGTATGGATA CAGCGATGAA GCAGTGCGACAACATAGACA 951 AGGGCAACCT TGA

This encodes a protein having amino acid sequence <SEQ ID 84; ORF15ng>:

1 MRARLLIPIL FSVFILSACG TLTGIPSHGG GKRFAVEQEL VAASARAAVK 51DMDLQALHGR KVALYIATMG DQGSGSLTGG RYSIDALIRG EYINSPAVRT 101DYTYPRYETT AETTSGGLTG LTTSLSTLNA PALSRTQSDG SGSRSSLGLN 151IGGMGDYRNE TLTTNPRDTA FLSHLVQTVF FLRGIDVVSP ANADTDVFIN 201IDVFGTIRNR TEMHLYNAET LKAQTKLEYF AVDRTNKKLL IKPKTNAFEA 251AYKENYALWM GPYKVSKGIK PTEGLMVDFS DIQPYGNHTG NSAPSVEADN 301SHEGYGYSDE AVRQHRQGQP *

The originally-identified partial strain B sequence (ORF15) shows 97.2%identity over a 213aa overlap with ORF15ng:

The complete strain B sequence (ORF15-1) and ORF15ng show 98.8% identityin 320 aa overlap:

Computer analysis of these amino acid sequences reveals an ILSAC motif(putative membrane lipoprotein lipid attachment site, as predicted bythe MOTIFS program).

Indicates a putative leader sequence, and it was predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

ORF15-1 (31.7 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 4A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 4B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for Western blot(FIG. 4C) and ELISA (positive result). These experiments confirm thatORFX-1 is a surface-exposed protein, and that it is a useful immunogen.

Example 11

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 85>:

1 ..GG.CAGCACA AAAAACAGGC GGTTGAACGG AAAAACCGTA TTTACGATGA 51  TGCCGGGTAT GATATTCGGC GTATTCACGG GCGCATTCTC CGCAAAATAT 101  ATCCCCGCGT TCGGGCTTCA AATTTTCTTC ATCCTGTTTT TAACCGCCGT 151  CGCATTCAAA ACACTGCATA CCGACCCTCA GACGGCATCC CGCCCGCTGC 201  CCGGACTGCC CrGACTGACT GCGGTTTCCA CACTGTTCGG CACAATGTCG 251  AGCTGGGTCG GCATAGGCGG CGGTTCACTT TCCGTCCCCT TCTTAATCCA 301  CTGCGGCTTC CCCGCCCATA AAGCCATCGG CACATCATCC GGCCTTGCCT 351  GGCCGATTGC ACTCTCCGGC GCAATATCGT ATCTGCTCAA CGGCCTGAAT 401  ATTGCAGGAT TGCCCGAAGG GTCACTGGGC TTCCTTTACC TGCCCGCCGT 451  CGCCGTCCTC AGCGCGGCAA CCATTGCCTT TGCCCCGCTC GGTGTCAAAA 501  CCGCCCACAA ACTTTCTTCT GCCAAACTCA AAAAATC.TT CGGCATTATG 551  TTGCTTTTGA TTGCCGGAAA AATGCTGTAC AACCTGCTTT AA

This corresponds to the amino acid sequence <SEQ ID 86; ORF17>:

1 ..GQHKKQAVNG KTVFTMMPGM IFGVFTGAFS AKYIPAFGLQ IFFILFLTAV 51  AFKTLHTDPQ TASRPLPGLP XLTAVSTLFG TMSSWVGIGG GSLSVPFLIH 101  CGFPAHKAIG TSSGLAWPIA LSGAISYLLN GLNIAGLPEG SLGFLYLPAV 151  AVLSAATIAF APLGVKTAHK LSSAKLKKSF GIMLLLIAGK MLYNLL*

Further work revealed the complete nucleotide sequence <SEQ ID 87>:

1 ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCCGTAG GCAGTGCGGC 51AGGTTTTATT GCCGGCCTGT TCGGCGTAGG CGGCGGCACG CTGATTGTCC 101CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA ACATCCTTAC 151GCGCAACACC TCGCCGTCGG CACATCCTTC GCCGTCATGG TCTTCACCGC 201CTTTTCCAGT ATGCTGGGGC AGCACAAAAA ACAGGCGGTC GACTGGAAAA 251CCGTATTTAC GATGATGCCG GGTATGATAT TCGGCGTATT CACGGGCGCA 301CTCTCCGCAA AATATATCCC CGCGTTCGGG CTTCAAATTT TCTTCATCCT 351GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGAC CCTCAGACGG 401CATCCCGCCC GCTGCCCGGA CTGCCCGGAC TGACTGCGGT TTCCACACTG 451TTCGGCACAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT CACTTTCCGT 501CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC ATCGGCACAT 551CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT ATCGTATCTG 601CTCAACGGCC TGAATATTGC AGGATTGCCC GAAGGGTCAC TGGGCTTCCT 651TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT GCCTTTGCCC 701CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA ACTCAAAAAA 751Tc.TTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC TGTACAACCT 801 GCTTTAA

This corresponds to the amino acid sequence <SEQ ID 88; ORF17-1>:

1 MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL DLQGLAQHPY 51AQHLAVGTSF AVMVFTAFSS MLGQHKKQAV DWKTVFTMMP GMIFGVFTGA 101LSAKYIPAFG LQIFFILFLT AVAFKTLHTD PQTASRPLPG LPGLTAVSTL 151FGTMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP IALSGAISYL 201LNGLNIAGLP EGSLGFLYLP AVAVLSAATI AFAPLGVKTA HKLSSAKLKK 251XFGIMLLLIA GKMLYNLL*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical H. influenzae Transmembrane Protein H10902(Accession number P44070)

ORF17 and HI0902 proteins show 28% aa identity in 192 aa overlap:

ORF17 3 HKKQAVNGKTVFTMMPGMIFGVFT-GAFSAKYIPAFGLQIF--FILFLTAVAFKTLHTDP 59HK   +  + V  + P ++  VF  G F  +       +IF   +++L      ++  D HI0902 72HKLGNIVWQAVRILAPVIMLSVFICGLFIGRLDREISAKIFACLVVYLATKMVLSIKKD- 130 ORF1760 QTASRPLPGLPXLTAVSTLFGTMSSWVGIGGGSLSVPFLIHCGFPAHKAIGTSSGLAWPI 119Q  ++ L  L  +     L G  SS  GIGGG   VPFL   G    +AIG+S+     + HI0902 131QVTTKSLTPLSSVIG-GILIGMASSAAGIGGGGFIVPFLTARGINIKQAIGSSAFCGMLL 189 ORF17120 ALSGAISYLLNGLNIAGLPEGSLGFLYLPAVAVLSAATIAFAPLGVXXXXXXXXXXXXXX 179 +SG  S++++G     +PE SLG++YLPAV  ++A +   + LG HI0902 190GISGMFSFIVSGWGNPLMPEYSLGYIYLPAVLGITATSFFTSKLGASATAKLPVSTLKKG 249 ORF17180 FGIMLLLIAGKM 191 F + L+++A  M HI0902 250 FALFLIVVAINM 261Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF17 shows 96.9% identity over a 196aa overlap with an ORF (ORF17a)from strain A of N. meningitidis:

The complete length ORF17a nucleotide sequence <SEQ ID 89> is:

1 ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCCGTAG GCAGTGCGGC 51AGGTTTTATT GCCGGCCTGT TCGGCGTAGG CGGCGGCACG CTGATTGTCC 101CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA ACATCCTTAC 151GCGCAACACC TCGCCGTCGG CACATCCTTC GCCGTCATGG TCTTCACCGC 201CTTTTCCAGT ATGCTGGGGC AGCACAAAAA ACAGGCGGTC GACTGGAAAA 251CCGTATTTAC GATGATGCCG GGTATGGTAT TCGGCGTATT CGCTGGCGCA 301CTCTCCGCAA AATATATCCC AGCGTTCGGG CTTCAAATTT TCTTCATCCT 351GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGAC CCTCAGACGG 401CATCCCGCCC GCTGCCCGGA CTGCCCGGAC TGACTGCGGT TTCCACACTG 451TTCGGCACAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT CACTTTCCGT 501CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC ATCGGCACAT 551CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT ATCGTATCTG 601CTCAACGGCC TGAATATTGC AGGATTGCCC GAAGGGTCAC TGGGCTTCCT 651TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT GCCTTTGCCC 701CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA ACTCAAAAAA 751TCCTTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC TGTACAACCT 801 GCTTTAA

This encodes a protein having amino acid sequence <SEQ ID 90>:

1 MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL DLQGLAQHPY 51AQHLAVGTSF AVMVFTAFSS MLGQHKKQAV DWKTVFTMMP GMVFGVFAGA 101LSAKYIPAFG LQIFFILFLT AVAFKTLHTD PQTASRPLPG LPGLTAVSTL 151FGTMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP IALSGAISYL 201LNGLNIAGLP EGSLGFLYLP AVAVLSAATI AFAPLGVKTA HKLSSAKLKK 251SFGIMLLLIA GKMLYNLL*

ORF17a and ORF17-1 show 98.9% identity in 268 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF17 shows 93.9% identity over a 196aa overlap with a predicted ORF(ORF17.ng) from N. gonorrhoeae:

An ORF17ng nucleotide sequence <SEQ ID 91> is predicted to encode aprotein having amino acid sequence <SEQ ID 92>:

1 MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL DLQGLAQHPY 51AQHLAVGTSF AVMVFTAFSS MLGQHKKQAV DWKTIFAMMP GMIFGVFAGA 101LSAKYIPAFG LQIFFILFLT AVAFKTLHTG RQTASRPLPG LPGLTAVSTL 151FGAMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP IALSGAISYL 201VNGLNIAGLP EGSLGFLYLP AVAVLSAATI AFAPLGVKTA HKLSSAKLKE 251SFGIMLLLIA GKMLYNLL*

Further work revealed the complete gonococcal DNA sequence <SEQ ID 93>:

1 ATGTGGCATT GGGACATTAT CTTAATCCTG CTTGCcgtag gcAGTGCGGC 51AGGTTTTATT GCCGGCCTGT Tcggtgtagg cggcgGTACG CTGATTGTCC 101CTGTCGTTTT ATGGGTGCTT GATTTGCAGG GTTTGGCACA ACATCCTTAC 151GCGCAACACC TCGCCGTCGG CAcaTccttc gcCGTCATGG TCTTCACCGC 201CTTTTCCAGT ATGTTGGGGC AGCACAAAAA ACAGGCGGTC GACTGGAAAA 251CCATATTTGC GATGATGCCG GGTATGATAT TCGGCGTATT CGCTGGCGCA 301CTCTCCGCAA AATATATCCC CGCGTTCGGG CTTCAAATTT TCTTCATCCT 351GTTTTTAACC GCCGTCGCAT TCAAAACACT GCATACCGGT CGTCAGACGG 401CATCCCGCCC GCTGCCCGGG CTGCCCGGAC TGACTGCGGT TTCCACACTG 451TTCGGCGCAA TGTCGAGCTG GGTCGGCATA GGCGGCGGTT CACTTTCCGT 501CCCCTTCTTA ATCCACTGCG GCTTCCCCGC CCATAAAGCC ATCGGCACAT 551CATCCGGCCT TGCCTGGCCG ATTGCACTCT CCGGCGCAAT ATCGTATCTG 601GTCAACGGTC TGAATATTGC AGGATTGCCC GAAGGGTCGC TGGGCTTCCT 651TTACCTGCCC GCCGTCGCCG TCCTCAGCGC GGCAACCATT GCCTTTGCCC 701CGCTCGGTGT CAAAACCGCC CACAAACTTT CTTCTGCCAA ACTCAAAGAA 751TCCTTCGGCA TTATGTTGCT TTTGATTGCC GGAAAAATGC TGTACAACCT 801 GCTTTAA

This corresponds to the amino acid sequence <SEQ ID 94; ORF17ng-1>:

1 MWHWDIILIL LAVGSAAGFI AGLFGVGGGT LIVPVVLWVL DLQGLAQHPY 51AQHLAVGTSF AVMVFTAFSS MLGQHKKQAV DWKTIFAMMP GMIFGVFAGA 101LSAKYIPAFG LQIFFILFLT AVAFKTLHTG RQTASRPLPG LPGLTAVSTL 151FGAMSSWVGI GGGSLSVPFL IHCGFPAHKA IGTSSGLAWP IALSGAISYL 201VNGLNIAGLP EGSLGFLYLP AVAVLSAATI AFAPLGVKTA HKLSSAKLKE 251SFGIMLLLIA GKMLYNLL*

ORF17ng-1 and ORF17-1 show 96.6% identity in 268 aa overlap:

In addition, ORF17ng-1 shows significant homology with a hypothetical H.influenzae protein:

sp|P44070|Y902_HAEIN HYPOTHETICAL PROTEIN HI0902 pir||G64015hypothetical protein HI0902 - Haemophilus influenzae (strain Rd KW20)gi|1573922 (U32772) H. influenzaepredicted coding region HI0902 [Haemophilus influenzae]Length = 264Score = 74 (34.9 bits), Expect = 1.6e−23, Sum P(2) = 1.6e−23Identities = 15/43 (34%), Positives = 23/43 (53%) Query: 55AVGTSFAVMVFTAFSSMLGQHKKQAVDWKTIFAMMPGMIFGVF 97A+GTSFA +V T   S    HK   + W+ +  + P ++  VF Sbjct: 52ALGTSFATIVITGIGSAQRHHKLGNIVWQAVRILAPVIMLSVF 94 Score =195 (91.9 bits), Expect = 1.6e−23, Sum P(2) = 1.6e−23 Identities =44/114 (38%), Positives = 65/114 (57%) Query: 150LFGAMSSWVGIGGGSLSVPFLIHCGFPAHKAIGTSSGLAWPIALSGAISYLVNGLNIAGL 209L G  SS  GIGGG   VPFL   G    +AIG+S+     + +SG  S++V+G     + Sbjct: 148LIGMASSAAGIGGGGFIVPFLTARGINIKQAIGSSAFCGMLLGISGMFSFIVSGWGNPLM 207 Query:210 PEGSLGFLYLPAVAVLSAATIAFAPLGVKTAFIKLSSAKLKESFGIMLLLIAGKM 263PE SLG++YLPAV  ++A +   + LG     KL  + LK+ F + L+++A  M Sbjct: 208PEYSLGYIYLPAVLGITATSFFTSKLGASATAKLPVSTLKKGFALFLIVVAINM 261

This analysis, including the homology with the hypothetical H.influenzae transmembrane protein, suggests that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 12

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 95>:

1 ..GGAAACGGAT GGCAGGCAGA CCCCGAACAT CCGCTGCTCG GGCTTTTTGC 51  CGTCAGTAAT GTATCGATGA CGCTTGCTTT TGTCGGAATA TGTGCGTTGG 101  TGCATTATTG CTTTTCGGGA ACGGTTCAAG TGTTTGTGTT TGCGGCACTG 151  CTCAAACTTT ATGCGCTGAA GCCGGTTTAT TGGTTCGTGT TGCAGTTTGT 201  GCTGATGGCG GTTGCCTATG TCCACCGCTG CGGTATAGAC CGGCAGCCGC 251  CGTCAACGTT CGGCGGCTCG CAGCTGCGAC TCGGCGGGTT GACGGCAGCG 301  TTGATGCAGG TCTCGGTACT GGTGCTGCTG CTTTCAGAAA TTGGAAGATA 351   A

This corresponds to the amino acid sequence <SEQ ID 96; ORF18>:

1 ..GNGWQADPEH PLLGLFAVSN VSMTLAFVGI CALVHYCFSG TVQVFVFAAL 51  LKLYALKPVY WFVLQFVLMA VAYVHRCGID RQPPSTFGGS QLRLGGLTAA 101  LMQVSVLVLL LSEIGR*

Further work revealed the complete nucleotide sequence <SEQ ID 97>:

1 ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGT ATGCGGCGGT 51TTTTCTGTTT CTGATATTCC GCGCAGGAAT GTTGCAATGG TTTTGGGCGA 101GTATTATGCT GTGGCTGGGC ATATCGGTTT TGGGGGCAAA GCTGATGCCC 151GGCATATGGG GAATGACCCG CGCCGCGCCC TTGTTCATCC CCCATTTTTA 201CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGCATTGG AACCGGAAAA 251CAGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCGCT GCTCGGGCTT 301TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG GAATATGTGC 351GTTGGTGCAT TATTGCTTTT CGGGAACGGT TCAAGTGTTT GTGTTTGCGG 401CACTGCTCAA ACTTTATGCG CTGAAGCCGG TTTATTGGTT CGTGTTGCAG 451TTTGTGCTGA TGGCGGTTGC CTATGTCCAC CGCTGCGGTA TAGACCGGCA 501GCCGCCGTCA ACGTTCGGCG GCTCGCAGCT GCGACTCGGC GGGTTGACGG 551CAGCGTTGAT GCAGGTCTCG GTACTGGTGC TGCTGCTTTC AGAAATTGGA 601 AGATAA

This corresponds to the amino acid sequence <SEQ ID 98; ORF18-1>:

1 MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIMLWLG ISVLGAKLMP 51 GIWGMTRAAPLFIPHFYLTL GSIFFFIGHW NRKTDGNGWQ ADPEHPLLGL 101 FAVSNVSMTL AFVGICALVHYCFSGTVQVF VFAALLKLYA LKPVYWFVLQ 151 FVLMAVAYVH RCGIDRQPPS TFGGSQLRLGGLTAALMQVS VLVLLLSEIG 201 R*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF18 shows 98.3% identity over a 116aa overlap with an ORF (ORF18a)from strain A of N. meningitidis:

The complete length ORF18a nucleotide sequence <SEQ ID 99> is:

1 ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGT ATGCGGCGGT 51 TTTTCTGTTTCTGATATTCC GCGCAGGAAT GTTGCAATGG TTTTGGGCGA 101 GTATTATGCT GTGGCTGGGCATATCGGTTT TGGGGGCAAA GCTGATGCCC 151 GGCATATGGG GAATGACCCG CGCCGCGCCCTTGTTCATCC CCCATTTTTA 201 CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGCATTGGAACCGGAAAA 251 CGGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCTCT GCTCGGGCTG301 TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG GAATATGTGC 351GTTGGTGCAT TATTGCTTTT CGNGAACGGT TCAAGTGTTT GTGTTTGCGG 401 CACTGCTCAAACTTTATGCG CTGAAGCCGG TTTATTGGTT CGTGTTGCAG 451 TTTGTGCTGA TGGCGGTTGCCTATGTCCAC CGCTGCGGTA TAGACCGGCA 501 GCCGCCGTCA ACGTTCGGCG GNTCGCAGCTGCGACTCGGC GGGTTGACGG 551 CAGCGTTGAT GCAGNTCTCG GTACTGGTGC TGCTGCTTTCAGAAATTGGA 601 AGATAA

This encodes a protein having amino acid sequence <SEQ ID 100>:

1 MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIMLWLG ISVLGAKLMP 51 GIWGMTRAAPLFIPHFYLTL GSIFFFIGHW NRKTDGNGWQ ADPEHPLLGL 101 FAVSNVSMTL AFVGICALVHYCFSXTVQVF VFAALLKLYA LKPVYWFVLQ 151 FVLMAVAYVH RCGIDRQPPS TFGGSQLRLGGLTAALMQXS VLVLLLSEIG 201 R*

ORF18a and ORF18-1 show 99.0% identity in 201 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF18 shows 93.1% identity over a 116aa overlap with a predicted ORF(ORF18.ng) from N. gonorrhoeae:

The complete length ORF18ng nucleotide sequence is <SEQ ID 101>:

1 ATGATTTTGC TGCATTTGGA TTTTTTGTCT GCCTTACTGt aTGCGGcggt 51 tttTctgTTTCTGATATTCC GCGCAGGAAT GTTGCAATGG TTTTGGGCGA 101 GTATTGCGTT GTGGCTCGGCATCTCGGTTT TAGGGGTAAA GCTGATGCCG 151 GGGATGTGGG GAATGACCCG CGCCGCGCCTTTGTTCATCC CCCATTTTTA 201 CCTGACTTTG GGCAGCATAT TTTTTTTCAT CGGGTATTGGAACCGGAAAA 251 CAGATGGAAA CGGATGGCAG GCAGACCCCG AACATCCGCT GCTCGGGCTT301 TTTGCCGTCA GTAATGTATC GATGACGCTT GCTTTTGTCG GAATATGTGC 351GTTGGTGCAT TATTGCTTTT CGGGAACGGT TCAAGTGTTT GTGTTTGCGG 401 CATTGCTCAAACTTTATGCG CTGAAGCCGG TTTATTGGTT CGTGTTGCAG 451 TTTGTATTGA TGGCGGttgCCTATGTCCAC CGCTGCGGTA TAGACCGGCA 501 GCCGCCGTCA ACGTTCGGCG GTTCGCAGCTGCGACTCGGC GTGTTGGCGG 551 CGATGTTGAT GCAGGTTGCG GTAACGGCGA TGCTGCTTGCCGAAATCGGC 601 AGATGA

This encodes a protein having amino acid sequence <SEQ ID 102>:

1 MILLHLDFLS ALLYAAVFLF LIFRAGMLQW FWASIALWLG ISVLGVKLMP 51 GMWGMTRAAPLFIPHFYLTL GSIFFFIGYW NRKTDGNGWQ ADPEHPLLGL 101 FAVSNVSMTL AFVGICALVHYCFSGTVQVF VFAALLKLYA LKPVYWFVLQ 151 FVLMAVAYVH RCGIDRQPPS TFGGSQLRLGVLAAMLMQVA VTAMLLAEIG 201 R*

This ORF18ng protein sequence shows 94.0% identity in 201 aa overlapwith ORF18-1:

Based on this analysis, including the presence of several putativetransmembrane domains in the gonococcal protein, it is predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 13

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 103>:

1 ATGAAAACCC CACTCCTCAA GCCTCTGCTN ATTACCTCGC TTCCCGTTTT 51 CGCCAGTGTTTTTACCGCCG CCTCCATCGT CTGGCAGCTA GGCGAACCCA 101 AGCTCGCCAT GCCCTTCGTACTCGGCATCA TCGCCGGCGG CCTTGTCGAT 151 TTGGACAACC NCNTGACCGG ACGGCTNAAAAACATCATCA CCACCGTCGC 201 CCTGTTCACC CTCTCCTCGC TCACGGCACA AAGCACCCTCGGCACAGGGC 251 TGCCCTTCAT CCTCGCCATG ACCCTGATGA CTT.CG.CTT CACCATTTTA301 GGCGCGGNCG ...

This corresponds to the amino acid sequence <SEQ ID 104; ORF 19>:

1 MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV LGIIAGGLVD 51 LDNXXTGRLKNIITTVALFT LSSLTAQSTL GTGLPFILAM TLMTXXFTIL 101 GAX...

Further work revealed the complete nucleotide sequence <SEQ ID 105>:

1 ATGAAAACCC CACTCCTCAA GCCTCTGCTC ATTACCTCGC TTCCCGTTTT 51 CGCCAGTGTTTTTACCGCCG CCTCCATCGT CTGGCAGCTA GGCGAACCCA 101 AGCTCGCCAT GCCCTTCGTACTCGGCATCA TCGCCGGCGG CCTTGTCGAT 151 TTGGACAACC GCCTGACCGG ACGGCTGAAAAACATCATCA CCACCGTCGC 201 CCTGTTCACC CTCTCCTCGC TCACGGCACA AAGCACCCTCGGCACAGGGC 251 TGCCCTTCAT CCTCGCCATG ACCCTGATGA CCTTCGGCTT CACCATTTTA301 GGCGCGGTCG GGCTCAAATA CCGCACCTTC GCCTTCGGTG CACTCGCCGT 351CGCCACCTAC ACCACACTTA CCTACACCCC CGAAACCTAC TGGCTGACCA 401 ACCCCTTCATGATTTTATGC GGCACCGTAC TGTACAGCAC CGCCATCCTC 451 CTGTTCCAAA TCGTCCTGCCCCACCGCCCC GTCCAAGAAA GCGTCGCCAA 501 CGCCTACGAC GCACTCGGCG GCTACCTCGAAGCCAAAGCC GACTTCTTCG 551 ACCCCGATGA GGCAGCCTGG ATAGGCAACC GCCACATCGACCTCGCCATG 601 AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT CCGCCCTGTT651 TTACCGCCTT CGCGGCAAAC ACCGCCACCC GCGCACCGCC AAAATGCTGC 701GTTACTACTT TGCCGCCCAA GACATACACG AACGCATCAG CTCCGCCCAC 751 GTCGATTATCAGGAAATGTC CGAAAAATTC AAAAACACCG ACATCATCTT 801 CCGCATCCAC CGCCTGCTCGAAATGCAGGG ACAAGCCTGC CGCAACACCG 851 CCCAAGCCCT GCGCGCAAGC AAAGACTACGTTTACAGCAA ACGCCTCGGC 901 CGCGCCATCG AAGGCTGCCG CCAATCGCTG CGCCTCCTTTCAGACAGCAA 951 CGACAGTCCC GACATCCGCC ACCTGCGCCG CCTTCTCGAC AACCTCGGCA1001 GCGTCGACCA GCAGTTCCGC CAACTCCAGC ACAACGGCCT GCAGGCAGAA 1051AACGACCGCA TGGGCGACAC CCGCATCGCC GCCCTCGAAA CCAGCAGCCT 1101 CAAAAACACCTGGCAGGCAA TCCGTCCGCA GCTAAACCTC GAATCAGGCG 1151 TATTCCGCCA TGCCGTCCGCCTGTCCCTCG TCGTTGCCGC CGCCTGCACC 1201 ATCGTCGAAG CCCTCAACCT CAACCTCGGCTACTGGATAC TACTGACCGC 1251 CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGCCGCGTCCGCC 1301 AGCGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC GCTCGTCCCC1351 TACTTCACCC CGTCTGTCGA AACCAAACTC TGGATTGTCA TCGCCAGTAC 1401CACCCTCTTT TTCATGACCC GCACCTACAA ATACAGTTTC TCCACCTTCT 1451 TCATTACCATTCAAGCCCTG ACCAGCCTCT CCCTCGCAGG TTTGGACGTA 1501 TACGCCGCCA TGCCCGTACGCATCATCGAC ACCATTATCG GCGCATCCCT 1551 TGCCTGGGCG GCAGTCAGCT ACCTGTGGCCAGACTGGAAA TACCTCACGC 1601 TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAACGGTGCCTATCTCGAA 1651 AAAATCACCG AACGCCTCAA AAGCGGCGAA ACCGGCGACG ACGTCGAATA1701 CCGCGCCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC CTCAGCAGCA 1751CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA CAGCCTGCAA 1801 CCCGGCTTTACCCTGCTCAA AACCGGCTAC GCCCTGACCG GCTACATCTC 1851 CGCCCTCGGC GCATACCGCAGCGAAATGCA CGAAGAATGC AGCCCCGACT 1901 TTACCGCACA GTTCCACCTC GCCGCCGAACACACCGCCCA CATCTTCCAA 1951 CACCTGCCCG AAACCGAACC CGACGACTTT CAGACAGCACTGGATACACT 2001 GCGCGGCGAA CTCGACACCC TCCGCACCCA CAGCAGCGGA ACACAAAGCC2051 ACATCCTCCT CCAACAGCTC CAACTCATCG CCCGACAGCT CGAACCCTAC 2101TACCGCGCCT ACCGCCAAAT TCCGCACAGG CAGCCCCAAA ATGCAGCCTG 2151 A

This corresponds to the amino acid sequence <SEQ ID 106; ORF19-1>:

1 MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV LGIIAGGLVD 51 LDNRLTGRLKNIITTVALFT LSSLTAQSTL GTGLPFILAM TLMTFGFTIL 101 GAVGLKYRTF AFGALAVATYTTLTYTPETY WLTNPFMILC GTVLYSTAIL 151 LFQIVLPHRP VQESVANAYD ALGGYLEAKADFFDPDEAAW IGNRHIDLAM 201 SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQDIHERISSAH 251 VDYQEMSEKF KNTDIIFRIH RLLEMQGQAC RNTAQALRAS KDYVYSKRLG301 RAIEGCRQSL RLLSDSNDSP DIRHLRRLLD NLGSVDQQFR QLQHNGLQAE 351NDRMGDTRIA ALETSSLKNT WQAIRPQLNL ESGVFRHAVR LSLVVAAACT 401 IVEALNLNLGYWILLTALFV CQPNYTATKS RVRQRIAGTV LGVIVGSLVP 451 YFTPSVETKL WIVIASTTLFFMTRTYKYSF STFFITIQAL TSLSLAGLDV 501 YAAMPVRIID TIIGASLAWA AVSYLWPDWKYLTLERTAAL AVCSNGAYLE 551 KITERLKSGE TGDDVEYRAT RRRAHEHTAA LSSTLSDMSSEPAKFADSLQ 601 PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL AAEHTAHIFQ651 HLPETEPDDF QTALDTLRGE LDTLRTHSSG TQSHILLQQL QLIARQLEPY 701YRAYRQIPHR QPQNAA*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Predicted Transmenbrane protein YHFK of H. influenzae(Accession Number P44289)

ORF19 and YHFK proteins show 45% aa identity in 97 aa overlap:

orf19 6 LKPLLITSLPVFASVFTAASIVWQLGEPKLAMPFVLGIIAGGLVDLDNXXTGRLKNIITT 65L   +I+++PVF +V  AA  +W       +MP +LGIIAGGLVDLDN  TGRLKN+  T YHFK 5LNAKVISTIPVFIAVNIAAVGIWFFDISSQSMPLILGIIAGGLVDLDNRLTGRLKNVFFT 64 orf19 66VALFTLSSLTAQSTLGTGLPFILAMTLMTXXFTILGA 102+  F++SS   Q  +G  + +I+ MT++T  FT++GA YHFK 65LIAFSISSFIVQLHIGKPIQYIVLMTVLTFIFTMIGA 101Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF19 shows 92.2% identity over a 102aa overlap with an ORF (ORF19a)from strain A of N. meningitidis:

The complete length ORF19a nucleotide sequence <SEQ ID 107> is:

1 ATGAAAACCC CACCCCTCAA GCCTCTGCTC ATTACCTCGC TTCCCGTTTT 51 CGCCAGTGTCTTTACCGCCG CCTCCATCGT CTGGCAGCTG GGCGAACCCA 101 AGCTCGCCAT GCCCTTCGTACTCGGCATCA TCGCTGGCGG CCTGGTCGAT 151 TTGGACAACC GCCTGACCGG ACGGCTGAAAAACATCATCG CCACCGTCGC 201 CCTGTTCACC CTCTCCTCAC TTGTCGCGCA AAGCACCCTCGGCACAGGTT 251 TGCCATTCAT CCTCGCCATG ACCCTGATGA CTTTCGGCTT TACCATCATG301 GGCGCGGTCG GGCTGAAATA CCGCACCTTC GCCTTCGGCG CACTCGCCGT 351CGCCACCTAC ACCACACTTA CCTACACCCC CGAAACCTAC TGGCTGACCA 401 ACCCCTTTATGATTCTGTGC GGAACCGTAC TGTACAGCAC CGCCATCATC 451 CTGTTCCAAA TCATCCTGCCCCACCGCCCC GTTCAAGAAA ACGTCGCCAA 501 CGCCTACGAA GCACTCGGCA GCTACCTCGAAGCCAAAGCC GACTTTTTCG 551 ATCCCGACGA AGCCGAATGG ATAGGCAACC GCCACATCGACCTCGCCATG 601 AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT CCGCCCTGTT651 TTACCGCCTT CGCGGCAAAC ACCGCCACCC GCGCACCGCC AAAATGCTGC 701GCTACTACTT CGCCGCCCAA GACATACACG AACGCATCAG CTCCGCCCAC 751 GTCGACTACCAAGAGATGTC CGAAAAATTC AAAAACACCG ACATCATCTT 801 CCGCATCCAC CGCCTGCTCGAAATGCAGGG ACAAGCCTGC CGCAACACCG 851 CCCAAGCCCT GCGCGCAAGC AAAGACTACGTTTACAGCAA ACGCCTCGGC 901 CGCGCCATCG AAGGCTGCCG CCAATCGCTG CGCCTCCTTTCAGACAGCAA 951 CGACAATCCC GACATCCGCC ACCTGCGCCG CCTTCTCGAC AACCTCGGCA1001 GCGTCGACCA GCAGTTCCGC CAACTCCAGC ACAACGGCCT GCAGGCAGAA 1051AACGACCGCA TGGGCGACAC CCGCATCGCC GCCCTCGAAA CCGGCAGCCT 1101 CAAAAACACCTGGCAGGCAA TCCGTCCGCA GCTAAACCTC GAATCAGGCG 1151 TATTCCGCCA TGCCGTCCGCCTGTCCCTTG TCGTTGCCGC CGCCTGCACC 1201 ATCGTCGAAG CCCTCAACCT CAACCTCGGCTACTGGATAC TACTGACCGC 1251 CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGCCGCGTCCGCC 1301 AGCGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC GCTCGTCCCC1351 TACTTTACCC CCTCCGTCGA AACCAAACTC TGGATCGTCA TCGCCAGTAC 1401CACCCTCTTT TTCATGACCC GCACCTACAA ATACAGCTTC TCGACATTTT 1451 TCATCACCATTCAAGCCCTG ACCAGCCTCT CCCTCGCAGG GTTGGACGTA 1501 TACGCCGCCA TGCCCGTACGCATCATCGAC ACCATTATCG GCGCATCCCT 1551 TGCCTGGGCG GCAGTCAGCT ACCTGTGGCCAGACTGGAAA TACCTCACGC 1601 TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAACGGCGCCTATCTCGAA 1651 AAAATCACCG AACGCCTCAA AAGCGGCGAA ACCGGCGACG ACGTCGAATA1701 CCGCGCCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC CTCAGCAGCA 1751CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA CAGCCTGCAA 1801 CCCGGCTTTACCCTGCTCAA AACCGGCTAC GCCCTGACCG GCTACATCTC 1851 CGCCCTCGGC GCATACCGCAGCGAAATGCA CGAAGAATGC AGCCCCGACT 1901 TTACCGCACA GTTCCACCTC GCCGCCGAACACACCGCCCA CATCTTCCAA 1951 CACCTGCCCG AAACCGAACC CGACGACTTT CAGACAGCACTGGATACACT 2001 GCGCGGCGAA CTCGACACCC TCCGCACCCA CAGCAGCGGA ACACAAAGCC2051 ACATCCTCCT CCAACAGCTC CAACTCATCG CCCGGCAGCT CGAACCCTAC 2101TACCGCGCCT ACCGACAAAT TCCGCACAGG CAGCCCCAAA ACGCAGCCTG 2151 A

This encodes a protein having amino acid sequence <SEQ ID 108>:

1 MKTPPLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV LGIIAGGLVD 51 LDNRLTGRLKNIIATVALFT LSSLVAQSTL GTGLPFILAM TLMTFGFTIM 101 GAVGLKYRTF AFGALAVATYTTLTYTPETY WLTNPFMILC GTVLYSTAII 151 LFQIILPHRP VQENVANAYE ALGSYLEAKADFFDPDEAEW IGNRHIDLAM 201 SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQDIHERISSAH 251 VDYQEMSEKF KNTDIIFRIH RLLEMQGQAC RNTAQALRAS KDYVYSKRLG301 RAIEGCRQSL RLLSDSNDNP DIRHLRRLLD NLGSVDQQFR QLQHNGLQAE 351NDRMGDTRIA ALETGSLKNT WQAIRPQLNL ESGVFRHAVR LSLVVAAACT 401 IVEALNLNLGYWILLTALFV CQPNYTATKS RVRQRIAGTV LGVIVGSLVP 451 YFTPSVETKL WIVIASTTLFFMTRTYKYSF STFFITIQAL TSLSLAGLDV 501 YAAMPVRIID TIIGASLAWA AVSYLWPDWKYLTLERTAAL AVCSNGAYLE 551 KITERLKSGE TGDDVEYRAT RRRAHEHTAA LSSTLSDMSSEPAKFADSLQ 601 PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL AAEHTAHIFQ651 HLPETEPDDF QTALDTLRGE LDTLRTHSSG TQSHILLQQL QLIARQLEPY 701YRAYRQIPHR QPQNAA*

ORF19a and ORF19-1 show 98.3% identity in 716 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF19 shows 95.1% identity over a 102aa overlap with a predicted ORF(ORF19.ng) from N. gonorrhoeae:

An ORF19ng nucleotide sequence <SEQ ID 109> is predicted to encode aprotein having amino acid sequence <SEQ ID 110>:

1 MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV LGIIAGGLVD 51LDNRLTGRLK NIIATVALFT LSSLTAQSTL GTGLPFILAM TLMTFGFTIL 101 GAVGLKYRTFAFGALAVATY TTLTYTPETY WLTNPFMILC GTVLYSTAII 151 LFQIILPHRPVQESVANAYE ALGGYLEAKA DFFDPDEAAW IGNRHIDLAM 201 SNTGVITAFN QCRSALFYRLRGKHRHPRTA KMLRYYFAAQ DIHERISSAH 251 VDYQEMSEKF KNTDIIFRIR RLLEMQGQACRNTAQAIRSG KDYVYSKRLG 301 RAIEGCRQSL RLLSDGNDSP DIRHLSRLLD NLGSVDQQFRQLRHSDSPAE 351 NDRMGDTRIA ALETGSFKNT *

Further work revealed the complete nucleotide sequence <SEQ ID 111>:

1 ATGAAAACCC CACTCCTCAA GCCTCTGCTC ATTACCTCGC TTCCCGTTTT 51 CGCCAGTGTCTTTACCGCCG CCTCCATCGT CTGGCAGCTA GGCGAACCCA 101 AGCTCGCCAT GCCCTTCGTACTCGGCATCA TCGCCGGCGG CCTGGTCGAT 151 TTGGACAACC GCCTGACCGG ACGGCTGAAAAACATCATCG CCACCGTCGC 201 CCTGTTTACC CTCTCCTCGC TCACGGCGCA AAGCACCCTCGGCACAGGGC 251 TGCCCTTCAT CCTCGCCATG ACCCTGATGA CCTTCGGCTT TACCATTTTA301 GGCGCGGTCG GGCTGAAATA CCGCACCTTC GCCTTCGGCG CACTCGCCGT 351CGCCACCTAC ACCACGCTTA CCTACACCCC CGAAACCTAC TGGCTGACCA 401 ACCCCTTCATGATTTTATGC GGCACCGTAC TGTACAGCAC CGCCATCATC 451 CTGTTCCAAA TCATCCTGCCCCACCGCCCC GTCCAAGAAA GCGTCGCCAA 501 TGCCTACGAA GCACTCGGCG GCTACCTCGAAGCCAAAGCC GACTTCTTCG 551 ACCCCGATGA GGCAGCCTGG ATAGGCAACC GCCACATCGACCTCGCCATG 601 AGCAACACCG GCGTCATCAC CGCCTTCAAC CAATGCCGTT CCGCCCTGTT651 TTACCGTTTG CGCGGCAAAC ACCGCCACCC GCGCACCGCC AAAATGCTGC 701GCTACTACTT CGCCGCCCAA GACATCCACG AACGCATCAG CTCCGCCCAC 751 GTCGACTACCAAGAGATGTC CGAAAAATTC AAAAACACCG ACATCATCTT 801 CCGCATCCGC CGCCTGCTCGAAATGCAGGG GCAGGCGTGC CGCAACACCG 851 CCCAAGCCAT CCGGTCGGGC AAAGACTAcgtTTACAGCAA ACGCCTCGGA 901 CGCGCCATcg aaggctgCCG CCAGTCGCtg cgcctCCTTtcagacggcaA 951 CGACAGTCCC GACATCCGCC ACCTGAGccg CCTTCTCGAC AACCTCGgca1001 GCGTcgacca gcagtTCcgc caactCCGAC ACAgcgactC CCCCGCcgaa 1051Aacgaccgca tgggcgacaC CCGCATCGCC GCCCtcgaaa ccggcagctT 1101 caaaaaCAcctggcaggCAA TCCGTCCGCa gctgaaCCTC GAATCatgCG 1151 TATTCCGCCA TGCCGTCCGCCTGTCCCTCG TCGTTGCCGC CGCCTGCACC 1201 ATCGTCgaag cCCTCAACCT CAACCTCGGCTACTGGATAC TGCTGACCGC 1251 CCTTTTCGTC TGCCAACCCA ACTACACCGC CACCAAAAGCCGCGTGTACC 1301 AACGCATCGC CGGCACCGTA CTCGGCGTAA TCGTCGGCTC GCTCGTCCCC1351 TACTTCACCC CCTCCGTCGA AACCAAACTC TGGATTGTCA TCGCCGGTAC 1401CACCCTGTTC TTCATGACCC GCACCTACAA ATACAGTTTC TCCACCTTCT 1451 TCATCACCATTCAGGCACTG ACCAGCCTCT CCCTCGCAGG TTTGGACGTA 1501 TACGCCGCCA TGCCCGTGCGCATCATcgaC ACCATTATCG GCGCATCCCT 1551 TGCCTGGGCG GCGGTCAGCT ACCTGTGGCCAGACTGGAAA TACCTCACGC 1601 TCGAACGCAC CGCCGCCCTT GCCGTATGCA GCAGCGGCACATACCTCCAA 1651 AAAATTGCCG AACGCCTCAA AACCGGCGAA ACCGGCGACG ACATAGAATA1701 CCGCATCACC CGCCGCCGCG CCCACGAACA CACCGCCGCC CTCAGCAGCA 1751CCCTTTCCGA CATGAGCAGC GAACCCGCAA AATTCGCCGA CAGCCTGCAA 1801 CCCGGCTTTACCCTGCTCAA AACCGGCTAC GCCCTGACCG GCTACATCTC 1851 CGCCCTCGGC GCATACCGCAGCGAAATGCA CGAAGAATGC AGCCCCGACT 1901 TTACCGCACA GTTCCACCTT GCCGCCGAACACACCGCCCA CATCTTCCAA 1951 CACCTGCCCG ACATGGGACC CGACGACTTT CAGACGGCATTGGATACACT 2001 GCGCGGCGAA CTCGGCACCC TCCGCACCCG CAGCAGCGGA ACACAAAGCC2051 ACATCCTCCT CCAACAGCTC CAACTCATCG CccgGCAACT CGAACCCTAC 2101TACCGCGCCT ACCGACAAAT TCCGCACAGG CAGCCCCAAA ACGCAGCCTG 2151 A

This corresponds to the amino acid sequence <SEQ ID 112; ORF19ng-1>:

1 MKTPLLKPLL ITSLPVFASV FTAASIVWQL GEPKLAMPFV LGIIAGGLVD 51 LDNRLTGRLKNIIATVALFT LSSLTAQSTL GTGLPFILAM TLMTFGFTIL 101 GAVGLKYRTF AFGALAVATYTTLTYTPETY WLTNPFMILC GTVLYSTAII 151 LFQIILPHRP VQESVANAYE ALGGYLEAKADFFDPDEAAW IGNRHIDLAM 201 SNTGVITAFN QCRSALFYRL RGKHRHPRTA KMLRYYFAAQDIHERISSAH 251 VDYQEMSEKF KNTDIIFRIR RLLEMQGQAC RNTAQAIRSG KDYVYSKRLG301 RAIEGCRQSL RLLSDGNDSP DIRHLSRLLD NLGSVDQQFR QLRHSDSPAE 351NDRMGDTRIA ALETGSFKNT WQAIRPQLNL ESCVFRHAVR LSLVVAAACT 401 IVEALNLNLGYWILLTALFV CQPNYTATKS RVYQRIAGTV LGVIVGSLVP 451 YFTPSVETKL WIVIAGTTLFFMTRTYKYSF STFFITIQAL TSLSLAGLDV 501 YAAMPVRIID TIIGASLAWA AVSYLWPDWKYLTLERTAAL AVCSSGTYLQ 551 KIAERLKTGE TGDDIEYRIT RRRAHEHTAA LSSTLSDMSSEPAKFADSLQ 601 PGFTLLKTGY ALTGYISALG AYRSEMHEEC SPDFTAQFHL AAEHTAHIFQ651 HLPDMGPDDF QTALDTLRGE LGTLRTRSSG TQSHILLQQL QLIARQLEPY 701YRAYRQIPHR QPQNAA*

ORF19ng-1 and ORF19-1 show 95.5% identity in 716 aa overlap:

In addition, ORF19ng-1 shows significant homology to a hypotheticalgonococcal protein previously entered in the databases:

sp|O33369|YOR2_NEIGO HYPOTHETICAL 45.5 KD PROTEIN (ORF2) gnl|PID|e1154438(AJ002423) hypothetical protein [Neisseria gonorrh] Length = 417 Score =1512 (705.6 bits), Expect = 5.3e−203, P = 5.3e−203 Identities =301/326 (92%), Positives = 306/326 (93%) Query: 307RQSLRLLSDGNDSPDIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS 366RQSLRLLSDGNDS DIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS Sbjct: 1RQSLRLLSDGNDSXDIRHLSRLLDNLGSVDQQFRQLRHSDSPAENDRMGDTRIAALETGS 60 Query:367 FKNTWQAIRPQLNLESCVFRHAVRLSLVVAAACTIVEALNLNLGYWILLTALFVCQPNYT 426FKNTWQAIRPQLNLES VFRHAVRLSLVVAAACTIVEALNLNLGYWILLT LFVCQPNYT Sbjct: 61FKNTWQAIRPQLNLESGVFRHAVRLSLVVAAACTIVEALNLNLGYWILLTRLFVCQPNYT 120 Query:427 ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT 486ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT Sbjct: 121ATKSRVYQRIAGTVLGVIVGSLVPYFTPSVETKLWIVIAGTTLFFMTRTYKYSFSTFFIT 180 Query:487 IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG 546IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG Sbjct: 181IQALTSLSLAGLDVYAAMPVRIIDTIIGASLAWAAVSYLWPDWKYLTLERTAALAVCSSG 240 Query:547 TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFADSLQPGFTLL 606TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFAD+  P Sbjct: 241TYLQKIAERLKTGETGDDIEYRITRRRAHEHTAALSSTLSDMSSEPAKFADTCNPALPCS 300 Query:607 KTGYALTGYISALGAYRSEMHEECSP 632 K   ALTGYISALG   ++  +  +P Sbjct: 301KPATALTGYISALGHTAAKCTKNAAP 326

Based on this analysis, including the presence of several putativetransmembrane domains in the gonococcal protein (the first of which isalso seen in the meningococcal protein), and on homology with the YHFKprotein, it is predicted that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 14

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 113>:

1 ATGAATATGC TGGGAGCTTT GGCAAAAGTC GGCAGCCTGA CGATGGTGTC 51GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG GCATTCGGCG 101CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT GCCCAACCTG 151CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT TTGTGCCGAT 201TTTGGCGGAA TACAAGGAAA CGCGTTCAAA AGAGGCGG.C GAAGCCTTTA 251TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTTAT CGTTACCGCG 301CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG CACCCGAGTT 351TTGCCCAAGA TGCCGACAAA TTTCAGCTCT CCATCGATTT GCTGCGGATT 401ACGTTTCCTT ATATATTATT GATTTCCCTG TCTTCATTTG TCGGCTCGGT 451ACTCAATTCT TATCATAAGT TCGGCATTCC GGCGTTTACG CCAC.GTTTC 501TGAACGTGTC GTTTATCGTA TTCGCGCTGT TTTTCGTGCC GTATTTCGAT 551CCGCCCGTTA CCGCGCyGGC GTGGGCGGTC TTTGTCGGCG GCATTTTGCA 601ACTCGrmTTC CAACTGCCCT GGCTGGCGAA ACTGGGCTTT TTGAAACTGC 651CCAAACtGAG TTTCAAAGAT GCGGCGGTCA ACCGCGTGAT GAAACAGATG 701GCGCCTGCgA TTTTgGGCGT GAgCGTGGCG CAGGTTTCTT TGGTGATCAA 751CACGATTTTc GCGTCTTATC TGCAATCGGG CAGCGTTTCA TGGATGTATT 801ACGCCGACCG CATGATGGAG CTGCCCAGCG GCGTGCTGGG GGCGGCACTC 851GGTACGATTT TGCTGCCGAC TTTGTCCAAA CACTCGGCAA ACCaAGATAC 901GGaACAGTTT TCCGCCCTGC TCGACTGGGG TTTGCGCCTG TGCATGCtgc 951TGACGCTGCC GGCGgcGGTC GGACTGGCGG TGTTGTCGTT cCCgCtGGTG 1001GCGACGCTGT TTATGTACCG CGwATTTACG CTGTTTGACG CGCAGATGAC 1051GCAACACGCG CTGATTGCCT ATTCTTTCGG TTTAATCGGC TTAATCATGA 1101TTAAAGTGTT GGCACCCGGC TTCTATGCGC GGCAAAACAT CAAwAmGCCC 1151GTCAAAATCG CCATCTTCAC GCTCATCTGC mCGCAGTTGA TGAACCTTGs 1201CTTTAyCGGC CCACTrrAAC rCasTCGGAC TTTCGCTTGC CATCGGTCTG 1251GGCGCGTGTA TCAATGCCGG ATTGTTGTTT TACCTGTTGC GCAGACACGG 1301TATTTACCAA CCTGG.CAAG GGTTGGGCAG CGTTCTT.AG CAAAAATGCT 1351GcTCTCGCTC GCCGTGA

This corresponds to the amino acid sequence <SEQ ID 114; ORF20>:

1 MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA FFVAFKLPNL 51LRRVFAEGAF AQAFVPILAE YKETRSKEAX EAFIRHVAGM LSFVLVIVTA 101LGILAAPWVI YVSAPSFAQD ADKFQLSIDL LRITFPYILL ISLSSFVGSV 151LNSYHKFGIP AFTPXFLNVS FIVFALFFVP YFDPPVTAXA WAVFVGGILQ 201LXFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV SVAQVSLVIN 251TIFASYLQSG SVSWMYYADR MMELPSGVLG AALGTILLPT LSKHSANQDT 301EQFSALLDWG LRLCMLLTLP AAVGLAVLSF PLVATLFMYR XFTLFDAQMT 351QHALIAYSFG LIGLIMIKVL APGFYARQNI XXPVKIAIFT LICXQLMNLX 401FXGPLXXIGL SLAIGLGACI NAGLLFYLLR RHGIYQPXQG LGSVLXQKCC 451 SRSP*

These sequences were elaborated, and the complete DNA sequence <SEQ ID115> is:

1 ATGAATATGC TGGGAGCTTT GGCAAAAGTC GGCAGCCTGA CGATGGTGTC 51GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG GCATTCGGCG 101CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT GCCCAACCTG 151CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT TTGTGCCGAT 201TTTGGCGGAA TACAAGGAAA CGCGTTCAAA AGAGGCGGCG GAGGCTTTTA 251TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTTAT CGTTACCGCG 301CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG CACCCGGTTT 351TGCCCAAGAT GCCGACAAAT TTCAGCTCTC CATCGATTTG CTGCGGATTA 401CGTTTCCTTA TATATTATTG ATTTCCCTGT CTTCATTTGT CGGCTCGGTA 451CTCAATTCTT ATCATAAGTT CGGCATTCCG GCGTTTACGC CCACGTTTCT 501GAACGTGTCG TTTATCGTAT TCGCGCTGTT TTTCGTGCCG TATTTCGATC 551CGCCCGTTAC CGCGCTGGCG TGGGCGGTCT TTGTCGGCGG CATTTTGCAA 601CTCGGCTTCC AACTGCCCTG GCTGGCGAAA CTGGGCTTTT TGAAACTGCC 651CAAACTGAGT TTCAAAGATG CGGCGGTCAA CCGCGTGATG AAACAGATGG 701CGCCTGCGAT TTTGGGCGTG AGCGTGGCGC AGGTTTCTTT GGTGATCAAC 751ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT GGATGTATTA 801CGCCGACCGC ATGATGGAGC TGCCCAGCGG CGTGCTGGGG GCGGCACTCG 851GTACGATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA CCAAGATACG 901GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCCTGT GCATGCTGCT 951GACGCTGCCG GCGGCGGTCG GACTGGCGGT GTTGTCGTTC CCGCTGGTGG 1001CGACGCTGTT TATGTACCGC GAATTTACGC TGTTTGACGC GCAGATGACG 1051CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGCT TAATCATGAT 1101TAAAGTGTTG GCACCCGGCT TCTATGCGCG GCAAAACATC AAAACGCCCG 1151TCAAAATCGC CATCTTCACG CTCATCTGCA CGCAGTTGAT GAACCTTGCC 1201TTTATCGGCC CACTGAAACA CGTCGGACTT TCGCTTGCCA TCGGTCTGGG 1251CGCGTGTATC AATGCCGGAT TGTTGTTTTA CCTGTTGCGC AGACACGGTA 1301TTTACCAACC TGGCAAGGGT TGGGCAGCGT TCTTAGCAAA AATGCTGCTC 1351TCGCTCGCCG TGATGTGCGG CGGACTGTGG GCAGCGCAGG CTTACCTGCC 1401GTTTGAATGG GCGCACGCCG GCGGAATGCG GAAAGCGGGG CAGCTCTGCA 1451TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCACT GGCGGCTTTG 1501GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAACTGA

This corresponds to the amino acid sequence <SEQ ID 116; ORF20-1>:

1 MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA FFVAFKLPNL 51LRRVFAEGAF AQAFVPILAE YKETRSKEAA EAFIRHVAGM LSFVLVIVTA 101LGILAAPWVI YVSAPGFAQD ADKFQLSIDL LRITFPYILL ISLSSFVGSV 151LNSYHKFGIP AFTPTFLNVS FIVFALFFVP YFDPPVTALA WAVFVGGILQ 201LGFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV SVAQVSLVIN 251TIFASYLQSG SVSWMYYADR MMELPSGVLG AALGTILLPT LSKHSANQDT 301EQFSALLDWG LRLCMLLTLP AAVGLAVLSF PLVATLFMYR EFTLFDAQMT 351QHALIAYSFG LIGLIMIKVL APGFYARQNI KTPVKIAIFT LICTQLMNLA 401FIGPLKHVGL SLAIGLGACI NAGLLFYLLR RHGIYQPGKG WAAFLAKMLL 451SLAVMCGGLW AAQAYLPFEW AHAGGMRKAG QLCILIAVGG GLYFASLAAL 501GFRPRHFKRV EN*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the MviN Virulence Factor of S. typhimurium (AccessionNumber P37169)

ORF20 and MviN proteins show 63% aa identity in 440aa overlap:

Orf20 1 MNMLGALAKVGSLTMVSRVLGFVRDTVIARAFGAGMATDAFFVAFKLPNLLRRVFAEGAF 60MN+L +LA V S+TM SRVLGF RD ++AR FGAGMATDAFFVAFKLPNLLRR+FAEGAF MviN 14MNLLKSLAAVSSMTMFSRVLGFARDAIVARIFGAGMATDAFFVAFKLPNLLRRIFAEGAF 73 Orf20 61AQAFVPILAEYKETRSKEAXEAFIRHVAGMLSFVLVIVTALGILAAPWVIYVSAPSFAQD 120+QAFVPILAEYK  + +EA   F+ +V+G+L+  L +VT  G+LAAPWVI V+AP FA MviN 74SQAFVPILAEYKSKQGEEATRIFVAYVSGLLTLALAVVTVAGMLAAPWVIMVTAPGFADT 133 Orf20121 ADKFQLSIDLLRITFPYILLISLSSFVGSVLNSYHKFGIPAFTPXFLNVSFIVFALFFVP 180ADKF L+  LLRITFPYILLISL+S VG++LN++++F IPAF P FLN+S I FALF  P MviN 134ADKFALTTQLLRITFPYILLISLASLVGAILNTWNRFSIPAFAPTFLNISMIGFALFAAP 193 Orf20181 YFDPPVTAXAWAVFVGGILQLXFQLPWLAKLGFLKLPKLSFKDAAVNRVMKQMAPAILGV 240YF+PPV A AWAV VGG+LQL +QLP+L K+G L LP+++F+D    RV+KQM PAILGV MviN 194YFNPPVLALAWAVTVGGVLQLVYQLPYLKKIGMLVLPRINFRDTGAMRVVKQMGPAILGV 253 Orf20241 SVAQVSLVINTIFASYLQSGSVSWMYYADRMMELPSGVLGAALGTILLPTLSKHSANQDT 300SV+Q+SL+INTIFAS+L SGSVSWMYYADR+ME PSGVLG ALGTILLP+LSK  A+ + MviN 254SVSQISLIINTIFASFLASGSVSWMYYADRLMEFPSGVLGVALGTILLPSLSKSFASGNH 313 Orf20301 EQFSALLDWGLRLCMLLTLPAAVGLAVLSFPLVATLFMYRXFTLFDAQMTQHALIAYSFG 360 +++ L+DWGLRLC LL LP+AV L +L+ PL  +LF Y  FT FDA MTQ ALIAYS G MviN 314DEYCRLMDWGLRLCFLLALPSAVALGILAKPLTVSLFQYGKFTAFDAAMTQRALIAYSVG 373 Orf20361 LIGLIMIKVLAPGFYARQNIXXPVKIAIFTLICXQLMNLXFXXXXXXXXXXXXXXXXXCI 420LIGLI++KVLAPGFY+RQ+I  PVKIAI TLI  QLMNL F                 C+ MviN 374LIGLIVVKVLAPGFYSRQDIKTPVKIAIVTLIMTQLMNLAFIGPLKHAGLSLSIGLAACL 433 Orf20421 NAGLLFYLLRRHGIYQPXQG 440 NA LL++ LR+  I+ P  G MviN 434NASLLYWQLRKQNIFTPQPG 453Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF20 shows 93.5% identity over a 447aa overlap with an ORF (ORF20a)from strain A of N. meningitidis:

The complete length ORF20a nucleotide sequence <SEQ ID 117> is:

1 ATGAATATGC TGGGAGCTTT GGTAAAAGTC GGCAGCCTGA CGATGGTGTC 51GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGC GCATTCGGCG 101CAGGCATGGC GACGGATGCG TTCTTTGTCG CGTTCAAACT GCCCAACCTG 151CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT TTGTGCCGAT 201TTTGGCGGAA TATAAGGAAA CGCGTTCTAA AGAGGCGACG GAGGCTTTTA 251TCCGCCATGT GGCGGGGATG CTGTCGTTTG TACTGGTCAT CGTTACCGCG 301CTGGGCATAC TTGCCGCGCC TTGGGTGATT TATGTTTCCG CACCCGGTTT 351TGCCAAAGAT GCCGACAAAT TTCAGCTCTC TATCGATTTG CTGCGGATTA 401CGTTTCCTTA TATCTTATTG ATTTCACTTT CCTCTTTTGT CGGCTCGGTA 451CTCAATTCCT ATCATAAATT CAGCATTCCT GCGTTTACGC CCACGTTCCT 501GAACGTGTCG TTTATCGTAT TCGCGCTGTT TTTCGTGCCG TATTTCGATC 551CTCCCGTTAC CGCGCTGGCT TGGGCGGTTT TTGTCGGCGG CATTTTGCAA 601CTCGGCTTCC AACTGCCCTG GCTGGCGAAA CTGGGTTTTT TGAAACTGCC 651CAAACTGAGT TTCAAAGATG CGGCGGTCAA CCGCGTGATG AAACAGATGG 701CGCCTGCGAT TTTGGGCGTG AGCGTGGCGC AGATTTCTTT GGTGATCAAC 751ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT GGATGTATTA 801CGCCGACCGC ATGATGGAAC TGCCCGGCGG CGTGCTGGGG GCGGCACTCG 851GTACGATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA CCAAGATACG 901GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCNTGT GCATGCTGCT 951GACGCTGCCG GCGGCGGTCG GAATGGCGGT GTTGTCGTTC CCGCTGGTGG 1001CAACCTTGTT TATGTACCGA GAATTCACGC TGTTTGACGC GCAGATGACG 1051CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGTT TAATCATGAT 1101TAAAGTGTTG GCGCCCGGCT TTTATGCGCG GCAAAACATC AAAACGCCCG 1151TCAAAATCGC CATCTTCACG CTCATTTGCA CGCAGTTGAT GAACCTTGCC 1201TTTATCGGCC CACTGAAACA CGTCGGACTT TCGCTTGCCA TCGGTCTGGG 1251CGCGTGTATC AATGCCGGAT TGTTGTTTTA CCTGTTGCGC AGACACGGTA 1301TTTACCAACC TGGCAAGGGT TGGGCAGCGT TCTTGGCAAA AATGCTGCTC 1351TCGCTCGCCG TGATGGGAGG CGGCCTGTAT GCCGCCCAAA TCTGGCTGCC 1401GTTCGACTGG GCACACGCCG GCGGAATGCA AAAGGCCGCC CGGCTCTTCA 1451TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCACT GGCGGCTTTG 1501GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAGCTGA

This encodes a protein having amino acid sequence <SEQ ID 118>:

1 MNMLGALVKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA FFVAFKLPNL 51LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM LSFVLVIVTA 101LGILAAPWVI YVSAPGFAKD ADKFQLSIDL LRITFPYILL ISLSSFVGSV 151LNSYHKFSIP AFTPTFLNVS FIVFALFFVP YFDPPVTALA WAVFVGGILQ 201LGFQLPWLAK LGFLKLPKLS FKDAAVNRVM KQMAPAILGV SVAQISLVIN 251TIFASYLQSG SVSWMYYADR MMELPGGVLG AALGTILLPT LSKHSANQDT 301EQFSALLDWG LRXCMLLTLP AAVGMAVLSF PLVATLFMYR EFTLFDAQMT 351QHALIAYSFG LIGLIMIKVL APGFYARQNI KTPVKIAIFT LICTQLMNLA 401FIGPLKHVGL SLAIGLGACI NAGLLFYLLR RHGIYQPGKG WAAFLAKMLL 451SLAVMGGGLY AAQIWLPFDW AHAGGMQKAA RLFILIAVGG GLYFASLAAL 501GFRPRHFKRV ES*

ORF20a and ORF20-1 show 96.5% identity in 512 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF20 shows 92.1% identity over a 454aa overlap with a predicted ORF(ORF20ng) from N. gonorrhoeae:

An ORF20ng nucleotide sequence <SEQ ID 119> was predicted to encode aprotein having amino acid sequence <SEQ ID 120>:

1 MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA FFVAFKLPNL 51LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM LSFVLIVVTA 101LGILAAPWVI YVSAPGFTKD ADKFQLSISL LRITFPYILL ISLSSFVGSI 151LNSYHKFGIP AFTPTFLNIS FIVFALFFVP YFDPPVTALA WAVFVGGILQ 201LGFQLPWLAK LGFLKLPKLN FKDAAVNRVM KQMAPAILGV SVAQISLVIN 251TIFASYLQSG SVSWMYYADR MMELPGGVLG AALGTILLPT LSKHSANQDT 301EQFSALLDWG LRLCMLLTLP AAAGLAVLSF PLVATLFMYR EFTLFDAQMT 351QHALIAYSFG LIGLIMIKVL ASGFYARQNI KTPVKIAIFT LICTQLMNLA 401FIGPLKHAGL SLAIGLGACI NAGLLFFLFR KHGIYRPGQG LGQPSWRKCC 451 SRSP*

Further DNA sequence analysis revealed the following DNA sequence <SEQID 121>:

1 ATGAATATGC TTGGAGCTTT GGCAAAAGTC GGCAGCCTGA CGATGGTGTC 51GCGCGTTTTG GGATTTGTGC GCGATACGGT CATTGCGCGG GCATTCGGCG 101CGGGTATGGC GACGGATGCG TTTTTTGTCG CGTTCAAACT GCCCAACCTG 151CTTCGCCGCG TGTTTGCGGA GGGGGCGTTT GCCCAAGCGT TTGTGCCGAT 201TTTGGCGGAA TATAAGGAAA CGCGTTCTAA AGAGGCGAcg gAGGCTTTTA 251TCCGCCACGt tgcgggAatg CTGTCGTTTG TGCTGATcgt cGttacCGCG 301CTGGGCATAC TTGCCGCgcc tTGGGTGATT TATGTTtccg CgcccGGCTT 351TACCAAAGAC GCGGACAAGT TCCAACTTTC CATCAGCCTG CTGCGGATTA 401CGTTTCCTTA TATATTATTG ATTTCTTTGT CTTCTTTTGT CGGCTCGATA 451CTCAATTCCT ACCATAAGTT CGGCATTCCC GCGTTTACGC CCACGTTTTT 501AAACATCTCT TTTATCGTAT TCGCACTGTT TTTCGTGCCG TATTTCGATC 551CGCCCGTTAC CGCGCTGGCG TGGGCGGTTT TTGTCGGCGG TATTTTGCAG 601CTCGGTTTCC AACTGCCGTG GCTGGCGAAA CTGGGCTTTT TGAAACTGCC 651CAAACTGAAT TTCAAAGATG CGGCGGTCAA CCGCGTCATG AAACAGATGG 701CGCCTGCGAT TTTGGGCGTG agcgTGGCGC AAATTTCTTT GgttATCAAC 751ACGATTTTCG CGTCTTATCT GCAATCGGGC AGCGTTTCAT GGATGTatta 801cgCCGACCGC ATGATGGAGc tgcgccGGGG CGTGCTGGGG GCTGCACTCG 851GTACAATTTT GCTGCCGACT TTGTCCAAAC ACTCGGCAAA CCAAGATACG 901GAACAGTTTT CCGCCCTGCT CGACTGGGGT TTGCGCCTGT GCATGCTGCT 951GACGCTGCCG GCGGCGGccg GACTGGCGGT ATTGTCGTTC CCGCTGGTGG 1001CGACGCTGTT TATGTACCGA GAATTCACGC TGTTTGACGC ACAAATGACG 1051CAACACGCGC TGATTGCCTA TTCTTTCGGT TTAATCGGTT TAATTATGAT 1101TAAAGTGTTG GCATCCGGCT TTTATGCGCG GCAAAACATC AAAACGCCCG 1151TCAAAATCGC CATCTTCACG CTCATCTGCA CGCAGTTGAT GAACCTCGCC 1201TTTATCGGTC CGTTGAAACA CGCCGGGCTT TCGCTCGCCA TCGGCCTGGG 1251CGCGTGCATC AACGCCGGAT TGTTGTTCTT CCTGTTGCGC AAACACGGTA 1301TTTACCGGCC cggcaggggt tgggcggcgt TCTTGGCGAA AATGCTGCTC 1351GCGCTCGCCG TGATGTGCGG CGGACTGTGG GCGGCGCAGG CTTGCCTGCC 1401GTTCGAATGG GCGCACGCCG GCGGAATGCG GAAAGCGGGG CAGCTCTGCA 1451TCCTGATTGC CGTCGGCGGC GGACTGTATT TCGCATCTCT GGCGGCTTTG 1501GGCTTCCGTC CGCGCCATTT CAAACGCGTG GAAAGCTGA

This encodes the following amino acid sequence <SEQ ID 122; ORF20ng-1>:

1 MNMLGALAKV GSLTMVSRVL GFVRDTVIAR AFGAGMATDA FFVAFKLPNL 51LRRVFAEGAF AQAFVPILAE YKETRSKEAT EAFIRHVAGM LSFVLIVVTA 101LGILAAPWVI YVSAPGFTKD ADKFQLSISL LRITFPYILL ISLSSFVGSI 151LNSYHKFGIP AFTPTFLNIS FIVFALFFVP YFDPPVTALA WAVFVGGILQ 201LGFQLPWLAK LGFLKLPKLN FKDAAVNRVM KQMAPAILGV SVAQISLVIN 251TIFASYLQSG SVSWMYYADR MMELRRGVLG AALGTILLPT LSKHSANQDT 301EQFSALLDWG LRLCMLLTLP AAAGLAVLSF PLVATLFMYR EFTLFDAQMT 351QHALIAYSFG LIGLIMIKVL ASGFYARQNI KTPVKIAIFT LICTQLMNLA 401FIGPLKHAGL SLAIGLGACI NAGLLFFLLR KHGIYRPGRG WAAFLAKMLL 451ALAVMCGGLW AAQACLPFEW AHAGGMRKAG QLCILIAVGG GLYFASLAAL 501GFRPRHFKRV ES*

ORF20ng-1 and ORF20-1 show 95.7% identity in 512 aa overlap:

In addition, ORF20ng-1 shows significant homology with a virulencefactor of S. typhimurium:

sp|P37169|MVIN_SALTY VIRULENCE FACTOR MVIN pir||S40271 mviN protein -Salmonella typhimurium gi|438252 (Z26133) mviB gene product[Salmonella typhimurium] gnl|PID|d1005521 (D25292) ORF2 [Salmonella typhimurium] Length = 524 Score =1573 (750.1 bits), Expect = 1.1e−220, Sum P(2) = 1.1e−220 Identities =309/467 (66%), Positives = 368/467 (78%) Query: 1MNMLGALAKVGSLTMVSRVLGFVRDTVIARAFGAGMATDAFFVAFKLPNLLRRVFAEGAF 60MN+L +LA V S+TM SRVLGF RD ++AR FGAGMATDAFFVAFKLPNLLRR+FAEGAF Sbjct: 14MNLLKSLAAVSSMTMFSRVLGFARDAIVARIFGAGMATDAFFVAFKLPNLLRRIFAEGAF 73 Query:61 AQAFVPILAEYKETRSKEATEAFIRHVAGMLSFVLIVVTALGILAAPWVIYVSAPGFTKD 120+QAFVPILAEYK  + +EAT  F+ +V+G+L+  L VVT  G+LAAPWVI V+APGF Sbjct: 74SQAFVPILAEYKSKQGEEATRIFVAYVSGLLTLALAVVTVAGMLAAPWVIMVTAPGFADT 133 Query:121 ADKFQLSISLLRITFPYILLISLSSFVGSILNSYHKFGIPAFTPTFLNISFIVFALFFVP 180ADKF L+  LLRITFPYILLISL+S VG+ILN++++F IPAF PTFLNIS I FALF  P Sbjct: 134ADKFALTTQLLRITFPYILLISLASLVGAILNTWNRFSIPAFAPTFLNISMIGFALFAAP 193 Query:181 YFDPPVTALAWAVFVGGILQLGFQLPWLAKLGFLKLPKLNFKDAAVNRVMKQMAPAILGV 240YF+PPV ALAWAV VGG+LQL +QLP+L K+G L LP++NF+D    RV+KQM PAILGV Sbjct: 194YFNPPVLALAWAVTVGGVLQLVYQLPYLKKIGMLVLPRINFRDTGAMRVVKQMGPAILGV 253 Query:241 SVAQISLVINTIFASYLQSGSVSWMYYADRMMELRRGVLGAALGTILLPTLSKHSANQDT 300SV+QISL+INTIFAS+L SGSVSWMYYADR+ME   GVLG ALGTILLP+LSK  A+ + Sbjct: 254SVSQISLIINTIFASFLASGSVSWMYYADRLMEFFSGVLGVALGTILLPSLSKSFASGNH 313 Query:301 EQFSALLDWGLRLCMLLTLPAAAGLAVLSFPLVATLFMYREFTLFDAQMTQHALIAYSFG 360 +++ L+DWGLRLC LL LP+A  L +L+ PL  +LF Y +FT FDA MTQ ALIAYS G Sbjct: 314DEYCRLMDWGLRLCFLLALPSAVALGILAKPLTVSLFQYGKFTAFDAAMTQRALIAYSVG 373 Query:361 LIGLIMIKVLASGFYARQNIKTPVKIAIFTLICTQLMNLAFIGPLKHAGLSLAIGLGACI 420LIGLI++KVLA GFY+RQ+IKTPVKIAI TLI TQLMNLAFIGPLKHAGLSL+IGL AC+ Sbjct: 374LIGLIVVKVLAPGFYSRQDIKTPVKIAIVTLIMTQLMNLAFIGPLKHAGLSLSIGLAACL 433 Query:421 NAGLLFFLLRKHGIYRPGRGWXXXXXXXXXXXXVMCGGLWAAQACLP 467 NA LL++ LRK  I+P  GW            VM   L+     +P Sbjct: 434NASLLYWQLRKQNIFTPQPGWMWFLMRLIISVLVMAAVLFGVLHIMP 480 Score =70 (33.4 bits), Expect = 1.1e−220, Sum P(2) = 1.1e−220 Identities =14/41 (34%), Positives = 23/41 (56%) Query: 469EWAHAGGMRKAGQLCILIAVGGGLYFASLAALGFRPRHFKR 509 EW+    + +  +L  ++ G   YFA+LA LGF+ + F R Sbjct: 481EWSQGSMLWRLLRLMAVVIAGIAAYFAALAVLGFKVKEFVR 521

Based on this analysis, including the homology with a virulence factorfrom S. typhimurium, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 15

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 123>:

1 atGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG GCAGACCGGA 51GCAAGCCGTT tACGACGGCC CGGCCaTTAC CGAAGtCGCG TTGCTTGGCG 101AAGAATATGC CGGTATGCGC CCCTCGATGA AAGTCAAGGA AGGCGATGCC 151GTcAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGAATC CGGGCGTGGT 201GTTTACTGCG CCGGCTTCAG GcAAAATCGC CGCGATTCAC CGTGGCGAAA 251AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAArGCAA CGACGAAATC 301GAGTTTGAAC GCTACGCACC TGAAGCGCTG GCAAACTTAA GCGGCGAAGA 351AGTGCGCCGC AACCTGATCC AATCCGGTTT GTGGACTGCG CTGCGCACCC 401GTCCGTTCAG CAAAATTCCT GCCGTCGATG CCGAGCCGTT CGCCATCTTC 451GTCAATGCGA tGGACACCAA TCCG..

This corresponds to the amino acid sequence <SEQ ID 124; ORF22>:

1 MIKIKKGLNL PIAGRPEQAV YDGPAITEVA LLGEEYAGMR PSMKVKEGDA 51VKKGQVLFED KKNPGVVFTA PASGKIAAIH RGEKRVLQSV VIAVEXNDEI 101EFERYAPEAL ANLSGEEVRR NLIQSGLWTA LRTRPFSKIP AVDAEPFAIF 151 VNAMDTNP..

Further work revealed the complete nucleotide sequence <SEQ ID 125>:

1 ATGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG GCAGACCGGA 51GCAAGCCGTT TACGACGGCC CGGCCATTAC CGAAGTCGCG TTGCTTGGCG 101AAGAATATGC CGGTATGCGC CCCTCGATGA AAGTCAAGGA AGGCGATGCC 151GTCAAAAAAG GCCAAGTGCT GTTTGAAGAC AAAAAGAATC CGGGCGTGGT 201GTTTACTGCG CCGGCTTCAG GCAAAATCGC CGCGATTCAC CGTGGCGAAA 251AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAAGGCAA CGACGAAATC 301GAGTTTGAAC GCTACGCACC TGAAGCGCTG GCAAACTTAA GCGGCGAAGA 351AGTGCGCCGC AACCTGATCC AATCCGGTTT GTGGACTGCG CTGCGCACCC 401GTCCGTTCAG CAAAATTCCT GCCGTCGATG CCGAGCCGTT CGCCATCTTC 451GTCAATGCGA TGGACACCAA TCCGCTGGCT GCCGACCCTA CGGTCATTAT 501CAAAGAAGCC GCCGAGGATT TCAAACGCGG CCTGTTGGTA TTGAGCCGTT 551TGACCGAACG CAAAATCCAT GTTTGTAAGG CAGCTGGCGC AGACGTGCCG 601TCTGAAAATG CTGCCAACAT CGAAACACAT GAATTCGGCG GCCCGCATCC 651TGCCGGTTTG AGTGGCACGC ACATTCATTT CATCGAGCCG GTCGGCGCGA 701ATAAAACCGT GTGGACCATC AATTATCAAG ATGTAATTAC CATTGGCCGT 751TTGTTTGCAA CAGGCCGTCT GAACACCGAG CGCGTGATTG CCCTAGGTGG 801TTCTCAAGTC AACAAACCGC GCCTCTTGCG TACCGTTTTG GGTGCGAAAG 851TATCGCAAAT TACTGCGGGC GAATTGGTTG ACACAGACAA CCGCGTGATT 901TCCGGTTCGG TATTGAACGG CGCGATTACA CAAGGCGCGC ACGATTATTT 951GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC CGCAGCAAAG 1001AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC CATCACGCGT 1051ACAACCCTCG GCCATTTCCT GAAAAACAAA CTCTTCAAGT TCAACACAGC 1101CGTCAACGGC GGCGACCGCG CCATGGTGCC GATTGGTACT TACGAGCGCG 1151TGATGCCCTT GGATATCCTG CCCACCCTGC TTTTGCGCGA TTTAATCGTC 1201GGCGATACCG ACAGCGCGCA GGCATTGGGT TGCTTGGAAT TGGACGAAGA 1251AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATAC GAATACGGCC 1301CGCTGTTGCG CAAAGTGCTG GAAACCATTG AGAAGGAAGG CTGA

This corresponds to the amino acid sequence <SEQ ID 126; ORF22-1>:

1 MIKIKKGLNL PIAGRPEQAV YDGPAITEVA LLGEEYAGMR PSMKVKEGDA 51 VKKGQVLFEDKKNPGVVFTA PASGKIAAIH RGEKRVLQSV VIAVEGNDEI 101 EFERYAPEAL ANLSGEEVRRNLIQSGLWTA LRTRPFSKIP AVDAEPFAIF 151 VNAMDTNPLA ADPTVIIKEA AEDFKRGLLVLSRLTERKIH VCKAAGADVP 201 SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTINYQDVITIGR 251 LFATGRLNTE RVIALGGSQV NKPRLLRTVL GAKVSQITAG ELVDTDNRVI301 SGSVLNGAIT QGAHDYLGRY HNQISVIEEG RSKELFGWVA PQPDKYSITR 351TTLGHFLKNK LFKFNTAVNG GDRAMVPIGT YERVMPLDIL PTLLLRDLIV 401 GDTDSAQALGCLELDEEDLA LCSFVCPGKY EYGPLLRKVL ETIEKEG*

Further work identified the corresponding gene in strain A of N.meningitidis <SEQ ID 127>:

1 ATGATTAAAA TCAAAAAAGG TCTAAACCTG CCCATCGCGG GCAGACCGGA 51 GCAAGTCATTTATGACGGGC CCGTCATTAC CGAAGTCGCG TTGCTTGGCG 101 AAGAATATGC CGGTATGCGCCCCTNGATGA AAGTCAAGGA AGGCGATGCC 151 GTCAAAAAAG GCCAAGTGCT GTTTGAAGACAAAAAGNATC CGGGCGTGGT 201 GTTTACCGCG CCNGTTTCAG GCAAAATCGC CGCCATCCATCGCGGCGAAA 251 AGCGCGTACT TCAGTCGGTC GTGATTGCCG TTGAAGGCAA CGACGAAATC301 GAGTTCGAAC GCTACGCGCC CGAAGCGTTG GCAAACTTAA GCGGCGANGA 351ANTNNGNNGC AATCTGATCC AATCCGGTTT GTGGACTGCG CTGCGTANCC 401 GTCCGTTCAGCAAAATCCCT GCCGTCGATG CCGAGCCGTT CGCCATCTTC 451 GTCAATGCGA TGGACACCAATCCGCTNGCG GCAGACCCTG TGGTTGTGAT 501 CAAAGAAGCC GNCGANGATT TCAGACGANGTNTGCTGGTA TTGAGCCGTT 551 TGACCGAGCG TAAAATCCAT GTGTGTAAGG CAGCTGGCGCAGACGTGCCG 601 TCTGAAAATG CTGCCAACAT CGAAACACAT GAATTCGGCG GCCCGCATCC651 GGCCGGTTTG AGTGGCACGC ACATTCATTT CATTGAGCCG GTCGGTGCAA 701ACAAAACCGT TTGGACCATC AATTATCAAG ATGTAATTGC CATCGGACGT 751 TTGTTTGCAACAGGCCGTCT GAACACCGAG CGCGTGATTG CTTTGGGTGG 801 TTCTCAAGTC AACAAACCACGCCTCTTGCG TACCGTTTTG GGTGCGAAAG 851 TATCGCAAAT TACTGCGGGC GAATTGGTTGACGCAGACAA CCGCGTGATT 901 TCCGGTTCGG TATTGAACGG CGCGATTACA CAAGGCGCGCACGATTATTT 951 GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC CGCAGCAAAG1001 AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC CATCACGCGT 1051ACGACCCTCG GCCATTTCCT GAAAAACAAA CTCTTCAAGT TCACGACAGC 1101 CGTCAACGGTGGCGACCGCG CCATGGTGCC GATTGGTACT TACGAGCGCG 1151 TAATGCCGCT AGACATCCTGCCTACCCTGC TTTTGCGCGA TTTAATCGTC 1201 GGCGATACCG ACAGCGCGCA AGCATTGGGTTGCTTGGAAT TGGACGAAGA 1251 AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATACGAATANGGCC 1301 CGCTGTTGCG TAAGGTGCTG GAAACCNTTG AGAAGGAAGG CTGA

This encodes a protein having amino acid sequence <SEQ ID 128; ORF22a>:

1 MIKIKKGLNL PIAGRPEQVI YDGPVITEVA LLGEEYAGMR PXMKVKEGDA 51 VKKGQVLFEDKKXPGVVFTA PVSGKIAAIH RGEKRVLQSV VIAVEGNDEI 101 EFERYAPEAL ANLSGXEXXXNLIQSGLWTA LRXRPFSKIP AVDAEPFAIF 151 VNAMDTNPLA ADPVVVIKEA XXDFRRXXLVLSRLTERKIH VCKAAGADVP 201 SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTINYQDVIAIGR 251 LFATGRLNTE RVIALGGSQV NKPRLLRTVL GAKVSQITAG ELVDADNRVI301 SGSVLNGAIT QGAHDYLGRY HNQISVIEEG RSKELFGWVA PQPDKYSITR 351TTLGHFLKNK LFKFTTAVNG GDRAMVPIGT YERVMPLDIL PTLLLRDLIV 401 GDTDSAQALGCLELDEEDLA LCSFVCPGKY EXGPLLRKVL ETXEKEG*

The originally-identified partial strain B sequence (ORF22) shows 94.2%identity over a 158aa overlap with ORF22a:

The complete strain B sequence (ORF22-1) and ORF22a show 94.9% identityin 447 aa overlap:

Further work identified a partial gene sequence <SEQ ID 129> from N.gonorrhoeae, which encodes the following amino acid sequence <SEQ ID130; ORF22ng>:

1 MIKIKKGLNL PIAGRPEQVI YDGPAITEVA LLGEEYVGMR PSMKIKEGEA 51 VKKGQVLFEDKKNPGVVFTA PASGKIAAIH RGEKRVLQSV VIAVEGNDEI 101 EFERYVPEAL AKLSSEKVRRNLIQSGLWTA LRTRPFSKIP AVDAEPFAIF 151 VNAMDTNPLA ADPTVIIKEA AEDFKRGLLVLSRLTERKIH VCKAAGADVP 201 SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTINYQDVIAIGR 251 LFVTGRLNTE RVVALGGLQV NKPRLLRTVL GAKVSQLTAG ELVDADNRVI301 SGSVLNGAIA QGAHDYLGRY HN*

Further work identified complete gonococcal gene <SEQ ID 131>:

1 ATGATTAAAA TCAAAAAAGG TCTAAATCTG CCCATCGCGG GCAGACCGGA 51 GCAAGTCATTTATGACGGCC CGGCCATTAC CGAAGTCGCG TTGCTTGGCG 101 AAGAATATGT CGGCATGCGCCCCTCGATGA AAATCAAGGA AGGTGAAGCC 151 GTCAAAAAAG GCCAAGTGCT GTTTGAAGACAAAAAGAATC CGGGCGTAGT 201 ATTTACTGCG CCGGCTTCAG GCAAAATCGC CGCTATTCACCGTGGCGAAA 251 AGCGCGTACT TCAGTCAGTC GTGATTGCCG TTGAAGGCAA CGACGAAATC301 GAGTTCGAAC GCTACGTACC TGAAGCGCTG GCAAAATTGA GCAGCGAAAA 351AGTGCGCCGC AACCTGATTC AATCAGGCTT ATGGACTGCG CTTCGCACCC 401 GTCCGTTCAGCAAAATCCCT GCCGTAGATG CCGAGCCGTT CGCCATCTTC 451 GTCAATGCGA TGGACACCAATCCGCTGGCT GCCGACCCTA CGGTCATCAT 501 CAAAGAAGCC GCCGAAGACT TCAAACGCGGCCTGTTGGTA TTGAGCCGCC 551 TGACCGAACG TAAAATCCAT GTGTGTAAAG CAGCAGGCGCAGACGTGCCG 601 TCTGAAAATG CTGCCAATAT CGAAACACAT GAATTTGGCG GCCCGCATCC651 TGCCGGCTTG AGTGGCACGC ACATTCATTT CATCGAGCCA GTCGGCGCGA 701ATAAAACCGT GTGGACCATC AATTATCAAG ACGTGATTGC TATCGGACGT 751 TTGTTCGTAACAGGCCGTCT GAATACCGAG CGCGTGGTTG CCTTGGGCGG 801 CCTGCAAGTC AACAAACCGCGCCTCTTGCG TACCGTTTTG GGTGCGAAGG 851 TGTCTCAACT TACCGCCGGC GAATTGGTTGACGCGGACAA CCGCGTGATT 901 TCCGGTTCGG TATTGAACGG TGCGATTGCA CAAGGCGCGCATGATTATTT 951 GGGACGCTAC CACAATCAGA TTTCCGTTAT CGAAGAAGGC CGCAGCAAAG1001 AGCTGTTCGG CTGGGTTGCG CCGCAGCCGG ACAAATACTC CATCACGCGC 1051ACCACTCTCG GCCATTTCCT AAAAAACAAA CTCTTCAAGT TCACGACAGC 1101 CGTCAACGGCGGCGACCGCG CCATGGTACC GATCGGCACT TATGAGCGCG 1151 TAATGCCGTT GGACATCCTGCCTACCTTGC TTTTGCGCGA TTTAATCGTC 1201 GGCGATACCG ACAGCGCGCA GGCTTTGGGTTGCTTGGAAT TGGACGAAGA 1251 AGACCTCGCT TTGTGCAGCT TCGTCTGCCC GGGCAAATACGAATACGGCC 1301 CGCTGTTGCG CAAAGTGCTG GAAACCATTG AGAAGGAAGG CTGA

This encodes a protein having amino acid sequence <SEQ ID 132;ORF22ng-1>:

1 MIKIKKGLNL PIAGRPEQVI YDGPAITEVA LLGEEYVGMR PSMKIKEGEA 51 VKKGQVLFEDKKNPGVVFTA PASGKIAAIH RGEKRVLQSV VIAVEGNDEI 101 EFERYVPEAL AKLSSEKVRRNLIQSGLWTA LRTRPFSKIP AVDAEPFAIF 151 VNAMDTNPLA ADPTVIIKEA AEDFKRGLLVLSRLTERKIH VCKAAGADVP 201 SENAANIETH EFGGPHPAGL SGTHIHFIEP VGANKTVWTINYQDVIAIGR 251 LFVTGRLNTE RVVALGGLQV NKPRLLRTVL GAKVSQLTAG ELVDADNRVI301 SGSVLNGAIA QGAHDYLGRY HNQISVIEEG RSKELFGWVA PQPDKYSITR 351TTLGHFLKNK LFKFTTAVNG GDRAMVPIGT YERVMPLDIL PTLLLRDLIV 401 GDTDSAQALGCLELDEEDLA LCSFVCPGKY EYGPLLRKVL ETIEKEG*

The originally-identified partial strain B sequence (ORF22) shows 93.7%identity over a 158aa overlap with ORF22ng:

The complete sequences from strain B (ORF22-1) and gonococcus (ORF22ng)show 96.2% identity in 447 aa overlap:

Computer analysis of these sequences gave the following results:

Homology with 48 kDa Outer Membrane Protein of Actinobacilluspleuropneumoniae (Accession Number U24492).

ORF22 and this 48 kDa protein show 72% aa identity in 158aa overlap:

Orf22 1 MIKIKKGLNLPIAGRPEQAVYDGPAITEVALLGEEYAGMRPSMKVKEGDAVKKGQVLFED 60MI IKKGL+LPIAG P Q +++G  + EVA+LGEEY GMRPSMKV+EGD VKKGQVLFED 48kDa 1MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED 60 orf22 61KKNPGVVFTAPASGKIAAIHRGEKRVLQSVVIAVEXNDEIEFERYAPEALANLSGEEVRR 120KKNPGVVFTAPASG +  I+RGEKRVLQSVVI VE +++I F RY    LA+LS E+V++ 48kDa 61KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ 120 orf22121 NLIQSGLWTALRTRPFSKIPAVDAEPFAIFVNAMDTNP 158 NLI+SGLWTA RTRPFSK+PA+DAP +IFVNAMDTNP 48kDa 121 NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNP 158

ORF22a also shows homology to the 48 kDa Actinobacillus pleuropneumoniaeprotein:

gi|1185395 (U24492) 48 kDa outer membrane protein [Actinobacilluspleuropneumoniae] Length = 449 Score = 530 bits (1351), Expect = e−150Identities = 274/450 (60%), Positives = 323/450 (70%), Gaps = 4/450 (0%)Query: 1 MIKIKKGLNLPIAGRPEQVIYDGPVITEVALLGEEYAGMRPXMKVKEGDAVKKGQVLFED 60MI IKKGL+LPIAG P QVI++G  + EVA+LGEEY GMRP MKV+EGD VKKGQVLFED Sbjct: 1MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED 60 Query:61 KKXPGVVFTAPVSGKIAAIHRGEKRVLQSVVIAVEGNDEIEFERYAPEALANLSGXEXXX 120 KKPGVVFTAP SG +  I+RGEKRVLQSVVI VEG+++I F RY    LA+LS  + Sbjct: 61KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ 120 Query:121 NLIQSGLWTALRXRPFSKIPAVDAEPFAIFVNAMDTNPLAADPVVVIKEAXXDFRRXXLV 180NLI+SGLWTA R RPFSK+PA+DA P +IFVNAMDTNPLAADP VV+KE   DF+    V Sbjct: 121NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNPLAADPEVVLKEYETDFKDGLTV 180 Query:181 LSRL--TERKIHVCKAAGADVP-SENAANIETHEFGGPHPAGLSGTHIHFIEPVGANKTV 237L+RL   ++ +++CK A +++P S     I    F G HPAGL GTHIHF++PVGA K V Sbjct: 181LTRLFNGQKPVYLCKDADSNIPLSPAIEGITIKSFSGVHPAGLVGTHIHFVDPVGATKQV 240 Query:238 WTINYQDVIAIGRLFATGRLNTERVIALGGSQVNKPRLLRTVLGAKVSQITAGELVDADN 297 W+NYQDVIAIG+LF TG L T+R+I+L G QV  PRL+RT LGA +SQ+TA EL   +N Sbjct: 241WHLNYQDVIAIGKLFTTGELFTDRIISLAGPQVKNPRLVRTRLGANLSQLTANELNAGEN 300 Query:298 RVISGSVLNGAITQGAHDYLGRYHNQISVIEEGRSKELFGWVAPQPDKYSITRTTLGHFL 357RVISGSVL+GA   G  DYLGRY  Q+SV+ EGR KELFGW+ P  DK+SITRT LGHF Sbjct: 301RVISGSVLSGATAAGPVDYLGRYALQVSVLAEGREKELFGWIMPGSDKFSITRTVLGHFG 360 Query:358 KNKLFKFTTAVNGGDRAMVPIGTYERVMXXXXXXXXXXXXXXVGDTDSAQXXXXXXXXXX 417 KKLF FTTAV+GG+RAMVPIG YERVM               GDTDSAQ Sbjct: 361K-KLFNFTTAVHGGERAMVPIGAYERVMPLDIIPTLLLRDLAAGDTDSAQNLGCLELDEE 419 Query:418 XXXXXSFVCPGKYEXGPLLRKVLETXEKEG 447      ++VCPGK   GP+LR  LE  EKEG

ORF22ng-1 also shows homology with the OMP from A. pleuropneumoniae:

gi|1185395 (U24492) 48 kDa outer membrane protein [Actinobacilluspleuropneumoniae] Length = 449 Score = 555 bits (1414), Expect = e−157Identities = 284/450 (63%), Positives = 337/450 (74%), Gaps = 4/450 (0%)Query: 27 MIKIKKGLNLPIAGRPEQVIYDGPAITEVALLGEEYVGMRPSMKIKEGEAVKKGQVLFED86 MI IKKGL+LPIAG P QVI++G  + EVA+LGEEYVGMRPSMK++EG+ VKKGQVLFED Sbjct: 1MITIKKGLDLPIAGTPAQVIHNGNTVNEVAMLGEEYVGMRPSMKVREGDVVKKGQVLFED 60 Query:87 KKNPGVVFTAPASGKIAAIHRGEKRVLQSVVIAVEGNDEIEFERYVPEALAKLSSEKVRR 146KKNPGVVFTAPASG +  I+RGEKRVLQSVVI VEG+++I F RY    LA LS+E+V++ Sbjct: 61KKNPGVVFTAPASGTVVTINRGEKRVLQSVVIKVEGDEQITFTRYEAAQLASLSAEQVKQ 120 Query:147 NLIQSGLWTALRTRPFSKIPAVDAEPFAIFVNAMDTNPLAADPTVIIKEAAEDFKRGLLV 206NLI+SGLWTA RTRPFSK+PA+DA P +IFVNAMDTNPLAADP V++KE   DFK GL V Sbjct: 121NLIESGLWTAFRTRPFSKVPALDAIPSSIFVNAMDTNPLAADPEVVLKEYETDFKDGLTV 180 Query:207 LSRL--TERKIHVCKAAGADVP-SENAANIETHEFGGPHPAGLSGTHIHFIEPVGANKTV 263L+RL   ++ +++CK A +++P S     I    F G HPAGL GTHIHF++PVGA K V Sbjct: 181LTRLFNGQKPVYLCKDADSNIPLSPAIEGITIKSFSGVHPAGLVGTHIHFVDPVGATKQV 240 Query:264 WTINYQDVIAIGRLFVTGRLNTERVVALGGLQVNKPRLLRTVLGAKVSQLTAGELVDADN 323 W+NYQDVIAIG+LF TG L T+R+++L G QV  PRL+RT LGA +SQLTA EL   +N Sbjct: 241WHLNYQDVIAIGKLFTTGELFTDRIISLAGPQVKNPRLVRTRLGANLSQLTANELNAGEN 300 Query:324 RVISGSVLNGAIAQGAHDYLGRYHNQISVIEEGRSKELFGWVAPQPDKYSITRTTLGHFL 383RVISGSVL+GA A G  DYLGRY  Q+SV+ EGR KELFGW+ P  DK+SITRT LGHF Sbjct: 301RVISGSVLSGATAAGPVDYLGRYALQVSVLAEGREKELFGWIMPGSDKFSITRTVLGHFG 360 Query:384 KNKLFKFTTAVNGGDRAMVPIGTYERVMXXXXXXXXXXXXXXVGDTDSAQXXXXXXXXXX 443 KKLF FTTAV+GG+RAMVPIG YERVM               GDTDSAQ Sbjct: 361K-KLFNFTTAVHGGERAMVPIGAYERVMPLDIIPTLLLRDLAAGDTDSAQNLGCLELDEE 419 Query:444 XXXXXSFVCPGKYEYGPLLRKVLETIEKEG 473      ++VCPGK  YGP+LR  LE IEKEGSbjct: 420 DLALCTYVCPGKNNYGPMLRAALEKIEKEG 449

Based on this analysis, including the homology with the outer membraneprotein of Actinobacillus pleuropneumoniae, it was predicted that theseproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

ORF22-1 (35.4 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 5A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 5B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for ELISA(positive result) and FACS analysis (FIG. 5C). These experiments confirmthat ORF22-1 is a surface-exposed protein, and that it is a usefulimmunogen.

Example 16

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 133>:

1 ..GCGnCGnAAA TCATCCATCC CC..nACGTC GTAGGCCCTG AAGCCAACTG 51  GTTTTTTATG GTAGCCAGTA CGTTTGTGAT TGCTTTGATT GGTTATTTTG 101  TTACTGAAAA AATCGTCGAA CCGCAATTGG GCCCTTATCA ATCAGATTTG 151  TCACAAGAAG AAAAAGACAT TCGGCATTCC AATGAAATCA CGCCTTTGGA 201  ATATAAAGGA TTAATTTGGG CTGGCGTGGT GTTTGTTGCC TTATCCGCCC 251  TATTGGCTTG GAGCATCGTC CCTGCCGACG GTATTTTGCG TCATCCTGAA 301  ACAGGATTGG TTTCCGGTTC GCCGTTTTTA AAATCGATTG TTGTTTTTAT 351  TTTCTTGTTG TTTGCACTGC CGGGCATTGT TTATGGCCGG GTAACCCGAA 401  GTTTGCGCGG CGAACAGGAA GTCGTTAATG CGmyGGCCGA ATCGATGAGT 451  ACTCTGGsGC TTTmTTTGsw CAkcATCTTT TTTGCCGCAC AGTTTGTCGC 501  ATTTTTTAAT TGGACGAATA TTGGGCAATA TATTGCCGTT AAAGGGGCGA 551  CGTTCTTAAA AGAAGTCGGC TTGGGCGGCA GCGTGTTGTT TATCGGTTTT 601  ATTTTAATTT GTGCTTTTAT CAATCTGATG ATAGGCTCCG CCTCCGCGCA 651  ATGGGCGGTA ACTGCGCCGA TTTTCGTCCC TATGCTGATG TTGGCCGGCT 701  ACGCGCCCGA AGTCATTCAA GCCGCTTACC GCATCGGTGA TTCCGTTACC 751  AATATTATTA CGCCGATGAT GAGTTATTTC GGGCTGATTA TGGCGACGGT 801  GrkCmmmTAC AAAAAAGATG CGGGCGTGGG TaCGcTGATT wCTATGATGT 851  TGCCGTATTC CGCTTTCTTC TTGATTGCgT GGATTGCCTT ATTCTGCATT 901  TGGGTATTTg TTTTGGGCCT GCCCGTCGGT CCCGGCGCGC CCACATTCTA 951  TCCCGCACCT TAA

This corresponds to the amino acid sequence <SEQ ID 134; ORF12>:

1 ..AXXIIHPXXV VGPEANWFFM VASTFVIALI GYFVTEKIVE PQLGPYQSDL 51  SQEEKDIRHS NEITPLEYKG LIWAGVVFVA LSALLAWSIV PADGILRHPE 101  TGLVSGSPFL KSIVVFIFLL FALPGIVYGR VTRSLRGEQE VVNAXAESMS 151  TLXLXLXXIF FAAQFVAFFN WTNIGQYIAV KGATFLKEVG LGGSVLFIGF 201  ILICAFINLM IGSASAQWAV TAPIFVPMLM LAGYAPEVIQ AAYRIGDSVT 251  NIITPMMSYF GLIMATVXXY KKDAGVGTLI XMMLPYSAFF LIAWIALFCI 301  WVFVLGLPVG PGAPTFYPAP *

Further sequence analysis revealed the complete DNA sequence <SEQ ID135> to be:

1 ATGAGTCAAA CCGATACGCA ACGGGACGGA CGATTTTTAC GCACAGTCGA 51 ATGGCTGGGCAATATGTTGC CGCATCCGGT TACGCTTTTT ATTATTTTCA 101 TTGTGTTATT GCTGATTGCCTCTGCCGTCG GTGCGTATTT CGGACTATCC 151 GTCCCCGATC CGCGCCCTGT TGGTGCGAAAGGACGTGCCG ATGACGGTTT 201 GATTTACATT GTCAGCCTGC TCAATGCCGA CGGTTTTATCAAAATCCTGA 251 CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG AACGGTGTTG301 GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT TGATTTCCGC 351ATTAATGCGC TTATTGCTCA CAAAATCGCC ACGCAAACTC ACTACTTTTA 401 TGGTTGTTTTTACAGGGATT TTATCTAATA CCGCTTCTGA ATTGGGCTAT 451 GTCGTCCTAA TCCCTTTGTCCGCCATCATC TTTCATTCCC TCGGCCGCCA 501 TCCGCTTGCC GGTCTGGCTG CGGCTTTCGCCGGCGTTTCG GGCGGTTATT 551 CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGCAGGCATCACC 601 CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG GCCCTGAAGC651 CAACTGGTTT TTTATGGTAG CCAGTACGTT TGTGATTGCT TTGATTGGTT 701ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC TTATCAATCA 751 GATTTGTCACAAGAAGAAAA AGACATTCGG CATTCCAATG AAATCACGCC 801 TTTGGAATAT AAAGGATTAATTTGGGCTGG CGTGGTGTTT GTTGCCTTAT 851 CCGCCCTATT GGCTTGGAGC ATCGTCCCTGCCGACGGTAT TTTGCGTCAT 901 CCTGAAACAG GATTGGTTTC CGGTTCGCCG TTTTTAAAATCGATTGTTGT 951 TTTTATTTTC TTGTTGTTTG CACTGCCGGG CATTGTTTAT GGCCGGGTAA1001 CCCGAAGTTT GCGCGGCGAA CAGGAAGTCG TTAATGCGAT GGCCGAATCG 1051ATGAGTACTC TGGGGCTTTA TTTGGTCATC ATCTTTTTTG CCGCACAGTT 1101 TGTCGCATTTTTTAATTGGA CGAATATTGG GCAATATATT GCCGTTAAAG 1151 GGGCGACGTT CTTAAAAGAAGTCGGCTTGG GCGGCAGCGT GTTGTTTATC 1201 GGTTTTATTT TAATTTGTGC TTTTATCAATCTGATGATAG GCTCCGCCTC 1251 CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATGCTGATGTTGG 1301 CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT CGGTGATTCC1351 GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC TGATTATGGC 1401GACGGTGATC AAATACAAAA AAGATGCGGG CGTGGGTACG CTGATTTCTA 1451 TGATGTTGCCGTATTCCGCT TTCTTCTTGA TTGCGTGGAT TGCCTTATTC 1501 TGCATTTGGG TATTTGTTTTGGGCCTGCCC GTCGGTCCCG GCGCGCCCAC 1551 ATTCTATCCC GCACCTTAA

This corresponds to the amino acid sequence <SEQ ID 136; ORF12-1>:

1 MSQTDTQRDG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA SAVGAYFGLS 51 VPDPRPVGAKGRADDGLIYI VSLLNADGFI KILTHTVKNF TGFAPLGTVL 101 VSLLGVGIAE KSGLISALMRLLLTKSPRKL TTFMVVFTGI LSNTASELGY 151 VVLIPLSAII FHSLGRHPLA GLAAAFAGVSGGYSANLFLG TIDPLLAGIT 201 QQAAQIIHPD YVVGPEANWF FMVASTFVIA LIGYFVTEKIVEPQLGPYQS 251 DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS IVPADGILRH301 PETGLVSGSP FLKSIVVFIF LLFALPGIVY GRVTRSLRGE QEVVNAMAES 351MSTLGLYLVI IFFAAQFVAF FNWTNIGQYI AVKGATFLKE VGLGGSVLFI 401GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGYAPEV IQAAYRIGDS 451VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA FFLIAWIALF 501 CIWVFVLGLPVGPGAPTFYP AP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF12 shows 96.3% identity over a 320aa overlap with an ORF (ORF12a)from strain A of N. meningitidis:

The complete length ORF12a nucleotide sequence <SEQ ID 137> is:

1 ATGAGTCAAA CCGATACGCA ACGGGACGGA CGATTTTTAC GCACAGTCGA 51 ATGGCTGGGCAATATGTTGC CGCACCCGGT TACGCTTTTT ATTATTTTCA 101 TTGTGTTATT GCTGATTGCCTCTGCCGCCG GTGCGTATTT CGGACTATCC 151 GTCCCCGATC CGCGCCCTGT TGGTGCGAAAGGACGTGCCG ATGACGGTTT 201 GATTCACGTT GTCAGCCTGC TCGATGCTGA CGGTTTGATCAAAATCCTGA 251 CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG AACGGTGTTG301 GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT TGATTTCCGC 351ATTAATGCGC TTATTGCTCA CAAAATCTCC ACGCAAACTC ACTACTTTTA 401 TGGTTGTTTTTACAGGGATT TTATCTAATA CCGCTTCTGA ATTGGGCTAT 451 GTCGTCCTAA TCCCTTTGTCCGCCATCATC TTTCATTCCC TCGGCCGCCA 501 TCCGCTTGCC GGTCTGGCTG CGGCTTTCGCCGGCGTTTCG GGCGGTTATT 551 CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGCAGGCATCACC 601 CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG GCCCTGAAGC651 CAACTGGTTT TTTATGGTAG CCAGTACGTT TGTGATTGCT TTGATTGGTT 701ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC TTATCAATCA 751 GATTTGTCACAAGAAGAAAA AGACATTCGA CATTCCAATG AAATCACGCC 801 TTTGGAATAT AAAGGATTAATTTGGGCTGG CGTGGTGTTT GTTGCCTTAT 851 CCGCCCTATT GGCTTGGAGC ATCGTCCCTGCCGACGGTAT TTTGCGTCAT 901 CCTGAAACAG GATTGGTTTC CGGTTCGCCG TTTTTAAAATCAATTGTTGT 951 TTTTATTTTC TTGTTGTTTG CACTGCCGGG CATTGTTTAT GGCCGGGTAA1001 CCCGAAGTTT GCGCGGCGAA CAGGAAGTCG TTAATGCGAT GGCCGAATCG 1051ATGAGTACTC TGGGGCTTTA TTTGGTCATC ATCTTTTTTG CCGCACAGTT 1101 TGTCGCATTTTTTAATTGGA CGAATATTGG GCAATATATT GCCGTTAAAG 1151 GGGCGACGTT CTTAAAAGAAGTCGGCTTGG GCGGCAGCGT GTTGTTTATC 1201 GGTTTTATTT TAATTTGTGC TTTTATCAATCTGATGATAG GCTCCGCCTC 1251 CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATGCTGATGTTGG 1301 CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT CGGTGATTCC1351 GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC TGATTATGGC 1401GACGGTGATC AAATACAAAA AAGATGCGGG CGTGGGTACG CTGATTTCTA 1451 TGATGTTGCCGTATTCCGCT TTCTTCTTGA TTGCGTGGAT TGCCTTATTC 1501 TGCATTTGGG TATTTGTTTTGGGCCTGCCC GTCGGTCCCG GCGCGCCCAC 1551 ATTCTATCCC GCACCTTAA

This encodes a protein having amino acid sequence <SEQ ID 138>:

1 MSQTDTQRDG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA SAAGAYFGLS 51 VPDPRPVGAKGRADDGLIHV VSLLDADGLI KILTHTVKNF TGFAPLGTVL 101 VSLLGVGIAE KSGLISALMRLLLTKSPRKL TTFMVVFTGI LSNTASELGY 151 VVLIPLSAII FHSLGRHPLA GLAAAFAGVSGGYSANLFLG TIDPLLAGIT 201 QQAAQIIHPD YVVGPEANWF FMVASTFVIA LIGYFVTEKIVEPQLGPYQS 251 DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS IVPADGILRH301 PETGLVSGSP FLKSIVVFIF LLFALPGIVY GRVTRSLRGE QEVVNAMAES 351MSTLGLYLVI IFFAAQFVAF FNWTNIGQYI AVKGATFLKE VGLGGSVLFI 401GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGYAPEV IQAAYRIGDS 451VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA FFLIAWIALF 501 CIWVFVLGLPVGPGAPTFYP AP*

ORF12a and ORF12-1 show 99.0% identity in 522 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF12 shows 92.5% identity over a 320aa overlap with a predicted ORF(ORF12.ng) from N. gonorrhoeae:

The complete length ORF12ng nucleotide sequence <SEQ ID 139> is:

1 ATGAGTCAAA CCGACGCGCG TCGTAGCGGA CGATTTTTAC GCACAGTCGA 51 ATGGCTGGGCAATATGTTGC CGCACCCGGT TACGCTTTTT ATTATTTTCA 101 TTGTGTTATT GCTGATTGcctctgCCGTCG GTGCGTATTT CGGACTATCC 151 GTCCCCGATC CGCGTCCTGT TGGGGCGAAAGGACGTGCCG ATGACGGTTT 201 GATTCACGTT GTCAGCCTGC TCGATGCCGA CGGTTTGATCAAAATCCTGA 251 CGCATACCGT TAAAAATTTC ACCGGTTTCG CGCCGTTGGG AACGGTGTTG301 GTTTCTTTAT TGGGCGTGGG GATTGCGGAA AAATCGGGCT TGATTTCCGC 351ATTAATGCGC TTATTGCTCA CAAAATCCCC ACGCAAACTC ACTACTTTTA 401 TGGTTGTTTTTACAGGGATT TTATCCAATA CGGCTTCTGA ATTGGGCTAT 451 GTCGTCCTAA TCCCTTTGTCCGCCGTCATC TTTCATTCGC TCGGCCGCCA 501 TCCGCTTGCC GGTTTGGCTG CGGCTTTCGCCGGCGTTTCG GGCGGTTATT 551 CGGCCAATCT GTTCTTAGGC ACAATCGATC CGCTCTTGGCAGGCATCACC 601 CAACAGGCGG CGCAAATCAT CCATCCCGAC TACGTCGTAG GCCCTGAAGC651 CAACTGGTTT TTTATGGCAG CCAGTACGTT TGTGATTGCT TTGATTGGTT 701ATTTTGTTAC TGAAAAAATC GTCGAACCGC AATTGGGCCC TTATCAATCA 751 GATTTGTCACAAGAAGAAAA AGACATTCGG CATTCCAATG AAATCACGCC 801 TTTGGAATAT AAAGGATTAATTTGGGCAGG CGTGGTGTTT GTTGCCTTAT 851 CCGCCCTATT GGCTTGGAGC ATCGTCCCTGCCGACGGTAT TTTGCGTCAT 901 CCTGAAACAG GATTGGTTGC CGGTTCGCCG TTTTTAAAATCGATTGTTGT 951 TTTTATTTTC TTGTTGTTTG CGCTGCCGGG CATTGTTTAT GGCCGGATAA1001 CCCGAAGTTT GCGCGGCGAA CGGGAAGTCG TTAATGCGAT GGCCGAATCG 1051ATGAGTACTT TGGGACTTTA TTTGGTCATC ATCTTTTTTG CCGCACAGTT 1101 TGTCGCATTTTTTAATTGGA CGAATATTGG GCAATATATT GCCGTTAAAG 1151 GGGCGGTGTT CTTAAAAGAAGTCGGCTTGG GCGGCAGTGT GTTGTTTATC 1201 GGTTTTATTT TAATTTGTGC TTTTATCAATCTGATGATAG GCTCCGCCTC 1251 CGCGCAATGG GCGGTAACTG CGCCGATTTT CGTCCCTATGCTGATGTTGG 1301 CCGGCTACGC GCCCGAAGTC ATTCAAGCCG CTTACCGCAT CGGTGATTCC1351 GTTACCAATA TTATTACGCC GATGATGAGT TATTTCGGGC TGATTATGGC 1401GACGGTAATC AAATACAAAA AAGATGCGGG CGTAGGCACG CTGATTTCTA 1451 TGATGTTGCCGTATTCCGCT TTCTTCTTAA TTGCATGGAT CGCCTTATTC 1501 TGCATTTGGG TATTTGTTTTGGGTCTGCCC GTCGGTCCCG GCACACCCAC 1551 ATTCTATCCG GTGCCTTAA

This encodes a protein having amino acid sequence <SEQ ID 140>:

1 MSQTDARRSG RFLRTVEWLG NMLPHPVTLF IIFIVLLLIA SAVGAYFGLS 51 VPDPRPVGAKGRADDGLIHV VSLLDADGLI KILTHTVKNF TGFAPLGTVL 101 VSLLGVGIAE KSGLISALMRLLLTKSPRKL TTFMVVFTGI LSNTASELGY 151 VVLIPLSAVI FHSLGRHPLA GLAAAFAGVSGGYSANLFLG TIDPLLAGIT 201 QQAAQIIHPD YVVGPEANWF FMAASTFVIA LIGYFVTEKIVEPQLGPYQS 251 DLSQEEKDIR HSNEITPLEY KGLIWAGVVF VALSALLAWS IVPADGILRH301 PETGLVAGSP FLKSIVVFIF LLFALPGIVY GRITRSLRGE REVVNAMAES 351MSTLGLYLVI IFFAAQFVAF  FNWTNIGQYI AVKGAVFLKK FRLGGSVLFI 401GFILICAFIN LMIGSASAQW AVTAPIFVPM LMLAGNAPQV IQAAYRIGDS 451VTNIITPMMS YFGLIMATVI KYKKDAGVGT LISMMLPYSA FFLIAWIALF 501 CIWVFVLGLPVGPGTPTFYP VP*

ORF12ng shows 97.1% identity in 522 aa overlap with ORF12-1:

In addition, ORF12ng shows significant homology with a hypotheticalprotein from E. coli:

sp|P46133|YDAH_ECOLI HYPOTHETICAL 55.1 KD PROTEIN IN OGT-DBPA INTERGENICREGION >gi|1787597 (AE000231) hypothetical protein in ogt 5′region[Escherichia coli] Length = 510 Score = 329 bits (835), Expect = 2e−89Identities = 178/507 (35%), Positives = 281/507 (55%), Gaps = 15/507(2%) Query: 8RSGRFLRTVEWLGNMLPHPVTXXXXXXXXXXXASAVGAYFGLSVPDPRPVGAKGRADDGL 67+SG+    VE +GN +PHP              +A+ + FG+S  +P         D Sbjct: 13QSGKLYGWVERIGNKVPHPFLLFIYLIIVLMVTTAILSAFGVSAKNP--------TDGTP 64 Query:68 IHVVSLLDADGLIKILTHTVKNFTGFAPXXXXXXXXXXXXIAEKSGLISALMRLLLTKSP 127 + V+LL  +GL   L + +KNF+GFAP            +AE+ GL+ ALM  + + Sbjct: 65VVVKNLLSVEGLHWFLPNVIKNFSGFAPLGAILALVLGAGLAERVGLLPALMVKMASHVN 124 Query:128 RKLTTFMVVFTGILSNTASELGYVVLIPLSAVIFHSLGRHPLAGLAAAFAGVSGGYSANL 187 +  ++MV+F    S+ +S+   V++ P+ A+IF ++GRHP+AGL AA AGV  G++ANL Sbjct: 125ARYASYMVLFIAFFSHISSDAALVIMPPMGALIFLAVGRHPVAGLLAAIAGVGCGFTANL 184 Query:188 FLGTIDPLLAGITQQAAQIIHPDYVVGPEANWFFMAASTFVIALIGYFVTEKIVEPQLGP 247 + T D LL+GI+ +AA   +P   V    NW+FMA+S  V+ ++G  +T+KI+EP+LG Sbjct: 185LIVTTDVLLSGISTEAAAAFNPQMHVSVIDNWYFMASSVVVLTIVGGLITDKIIEPRLGQ 244 Query:248 YQSDLSQEEKDIRHSNEITPLEYKGLIWAGVVFVALSALLAWSIVPADGILRHPETGLVA 307 +Q+  ++ + +  S         GL  AGVV +   A +A  ++P +GILR P    V Sbjct: 245WQGNSDEKLQTLTESQRF------GLRIAGVVSLLFIAAIALMVIPQNGILRDPINHTVM 298 Query:308 GSPFLKSIVVFIFLLFALPGIVYGRITRSLRGEREVVNAMAESMSTLGLYLXXXXXXXXX 367 SPF+K IV  I L F +  + YG  TR++R + ++ + M E M  +  ++ Sbjct: 299PSPFIKGIVPLIILFFFVVSLAYGIATRTIRRQADLPHLMIEPMKEMAGFIVMVFPLAQF 358 Query:368 XXXXNWTNIGQYIAVKGAVFLKEVGLGGSVLFIGFILICAFINLMIGSASAQWAVTAPIF 427    NW+N+G++IAV     L+  GL G   F+G  L+ +F+ +I S SA W++ APIF Sbjct: 359VAMFNWSNMGKFIAVGLTDILESSGLSGIPAFVGLALLSSFLCMFIASGSAIWSILAPIF 418 Query:428 VPMLMLAGYAPEVIQAAYRIGDSVTNIITPMMSYFGLIMATVIKYKKDAGVGTLISMMLP 487 VPMML G+ P   Q  +RI DS    + P+  +  L +  + +YK DA +GT  S++LP Sbjct: 419VPMFMLLGFHPAFAQILFRIADSSVLPLAPVSPFVPLFLGFLQRYKPDAKLGTYYSLVLP 478 Query:488 YSAFFLIAWIALFCIWVFVLGLPVGPG 514 Y   FL+ W+ +   W +++GLP+GPG Sbjct:479 YPLIFLVVWLLMLLAW-YLVGLPIGPG 504

Based on this analysis, including the presence of several putativetransmembrane domains and the predicted actinin-type actin-bindingdomain signature (shown in bold) in the gonococcal protein, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 17

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 141>:

1 ..ACAGCCGGCG CAGCAGGTTn CnCGGTCTTC GTTTTCGTAA CGGACAGTCA 51  GGTGGAGGTG TTCGGGAACA TCCAGACCGC AGTGGAAACA GGTTTTTTTC 101  ATGGCATTTC GGTTTCGTCT GTGTTTGGTG CGGCGGCACA AGACTCGGCA 151  ATgGCTTCGC GCAGTGCGTC TATACCGGTA TTTTCAGCAA CGGAAATGCG 201  GACGGcGgCA ATTTTTCCCG CAGCGTCGCG CCATATGCCC GTGTTTTgTT 251  CTTCAGACGG CAGCAGGTCG GTTTTGTTGT ACACCTTgAT GCACGGAaTA 301  TCGCCGGCAT GGATTTCTTG CAGTACGTTT TCCACGTCTT CAATCTGCTG 351  TCCGCTGTTC GGAGCGGCGG CATCGACGAC GTGCAGCAGC ACATCgGcTT 401  gCGCGGTTTC TTCCAGCGTG GCgGAAAAGG CGGAAATCAG TTTgTGCGGC 451  agATyGCTnA CGAATCCGAC GGTATCGGTC AGGATAATGC TGCATTCGGG 501   ACT..

This corresponds to the amino acid sequence <SEQ ID 142; ORF14>:

1 ..TAGAAGXXVF VFVTDSQVEV FGNIQTAVET GFFHGISVSS VFGAAAQDSA 51  MASRSASIPV FSATEMRTAA IFPAASRHMP VFCSSDGSRS VLLYTLMHGI 101  SPAWISCSTF STSSICCPLF GAAASTTCSS TSACAVSSSV AEKAEISLCG 151  RXLTNPTVSV RIMLHSG..

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF From N. meningitidis (Strain A)

ORF14 shows 94.0% identity over a 167aa overlap with an ORF (ORF14a)from strain A of N. meningitidis:

The complete length ORF14a nucleotide sequence <SEQ ID 143> is:

1 ATGGAGGATT TGCAGGAAAT CGGGTTCGAT GTCGCCGCCG TAAAGGTAGG 51 TCGGCAGCGCGAACATCATC GTCTGCATCA TCCCCAGCCC GGCAACGGCG 101 AGGCGGACGA TGTATTGTTTGCGTTCTTTT TGGTTGGCGG CTTCGATTTT 151 TTGCGCGTCA TAGGGTGCGG CGGTGTAGCCTATCTGCCTG ATTTTCAACA 201 GAATGTCGGA AAGGCGGATT TTGCCGTCGT CCCAGACGACGCGGCAGCGG 251 TGCGTGCTGT AATTGAGGTC GATGCGGACG ATGCCGTCTG TACGCAAAAG301 CTGCTGTTCG ATCAGCCAGA CGCAGGCGGC GCAGGTGATG CCGCCGAGCA 351TTAAAACCGC CTCGCGCGTG CCGCCGTGGG TTTCCACAAA GTCGGACTGG 401 ACTTCGGGCAGGTCGTACAG GCGGATTTGG TCGAGGATTT CTTGGGGCGG 451 CAGCTCGGTT TTTTGCGCGTCGGCGGTGCG TTGTTTGTAA TAACTGCCCA 501 AGCCCGCGTC AATAATGCTT TGTGCGACTGCCTGACAACC GGCGCAGCAG 551 GTTTCGCGGT CTTCGTTTTC GTAACGGACG GTCAGATGCAGGTTTTCGGG 601 AACGTCCAGC CCGCAGTGGA AACAGGTTTT TTTCATGGCA TTTCGGTTTC651 GTCTGTGTTT GGTGCGGCGG CACAATACTC GGCAATGGCT TCGCGCAGTG 701CGTCTATACC GGTATTTTCA GCAACGGAAA TGCGGACGGC GGCAATTTTT 751 CCCGCAGCGTCGCGCCATAT GCCCGTGTTT TGTTCTTCAG ACGGCAGCAG 801 GTCGGTTTTG TTGTACACCTTGATGCACGG AATATCGCCG GCATGGATTT 851 CTTGCAGTAC GTTTTCCACG TCTTCAATCTGCTGTCCGCT GTTCGGAGCG 901 GCGGCATCGA CGACGTGCAG CAGCACATCG GCTTGCGCGGTTTCTTCCAG 951 CGTGGCGGAA AAGGCGGAAA TCAGTTTGTG CGGCAGATCG CTGACGAATC1001 CGACGGTATC GGTCAGGATA ATGCTGCATT CGGGACTGAT GTACAGCCGC 1051CGCGCCGTCG TGTCGAGTGT GGCGAAAAGC TGGTCTTTCG CATATATGCC 1101 CGACTTGGTCAGCCGGTTGA ACAGACTGGA TTTGCCGACA TTGGTATAG

This encodes a protein having amino acid sequence <SEQ ID 144>:

1 MEDLQEIGFD VAAVKVGRQR EHHRLHHPQP GNGEADDVLF AFFLVGGFDF 51 LRVIGCGGVAYLPDFQQNVG KADFAVVPDD AAAVRAVIEV DADDAVCTQK 101 LLFDQPDAGG AGDAAEH*NRLARAAVGFHK VGLDFGQVVQ ADLVEDFLGR 151 QLGFLRVGGA LFVITAQARV NNALCDCLTTGAAGFAVFVF VTDGQMQVFG 201 NVQPAVETGF FHGISVSSVF GAAAQYSAMA SRSASIPVFSATEMRTAAIF 251 PAASRHMPVF CSSDGSRSVL LYTLMHGISP AWISCSTFST SSICCPLFGA301 AASTTCSSTS ACAVSSSVAE KAEISLCGRS LTNPTVSVRI MLHSGLMYSR 351RAVVSSVAKS WSFAYMPDLV SRLNRLDLPT LV*

It should be noted that this sequence includes a stop codon at position118.

Homology with a Predicted ORF from N. gonorrhoeae

ORF14 shows 89.8% identity over a 167aa overlap with a predicted ORF(ORF14.ng) from N. gonorrhoeae:

The complete length ORF14ng nucleotide sequence <SEQ ID 145> ispredicted to encode a protein having amino acid sequence <SEQ ID 146>:

1 MEDLQEIGFD VAAVKVGRQR EHHRLHHTQS GNGKADDVLF AFFLVGGFDF 51 LRVIGCGGVACLPDFQQNVG EADFAVVPDD AAAVRAVIEV DADDAVCAQK 101 LLFDQPDAGG AGNAAEHQHCFVRAIMGFHK VGLDFGQVVQ ADLVEDFLGR 151 QFGFFRVGGA SFVITAQAGI DDALCDCLTADAAGFAVFAF VADGQMQVFG 201 NVQPAVETGF FHGISVSSVF GAAAQYSAMA SRSASIPVFSATEMRTAAIF 251 PAASRHMPVF CSSDGSRSVL LYTLMHGISW AWISCSTFST SSICCPLFRA301 AASTTCSSTS ACTVSSKVAE KAEISLCGRS LTNPTVSVRI MLHAGLMYSR 351RAVVSRVAKS WSFAYMPDLV SRLNRLDLPT LV*

Based on the putative transmembrane domain in the gonococcal protein, itis predicted that the proteins from N. meningitidis and N. gonorrhoeae,and their epitopes, could be useful antigens for vaccines ordiagnostics, or for raising antibodies.

Example 18

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 147>:

1 ..GGCCATTACT CCGACCGCAC TTGGAAGCCG CGTTTGGNCG GCCGCCGTCT 51  GCCGTATCTG CTTTATGGCA CGCTGATTGC GGTTATTGTG ATGATTTTGA 101  TGCCGAACTC GGGCAGCTTC GGTTTCGGCT ATGCGTCGCT GGCGGCTTTG 151  TCGTTCGGCG CGCTGATGAT TGCGCTGTTA GACGTGTCGT CAAATATGGC 201  GATGCAGCCG TTTAAGATGA TGGTCGGCGA CATGGTCAAC GAGGAGCAGA 251  AAA.NTACGC CTACGGGATT CAAAGTTTCT TAGCAAATAC GGGCGCGGTC 301  GTGGCGGCGA TTCTGCCGTT TGTGTTTGCG TATATCGGTT TGGCGAACAC 351  CGCCGANAAA GGCGTTGTGC CGCAGACCGT GGTCGTGGCG TTTTATGTGG 401  GTGCGGCGTT GCTGGTGATT ACCAGCGCGT TCACGATTTT CAAAGTGAAG 451  GAATACGANC CGGAAACCTA CGCCCGTTAC CACGGCATCG ATGTCGCCGC 501  GAATCAGGAA AAAGCCAACT GGATCGCACT CTTAAAA.CC GCGC..

This corresponds to the amino acid sequence <SEQ ID 148; ORF16>:

1 ..GHYSDRTWKP RLXGRRLPYL LYGTLIAVIV MILMPNSGSF GFGYASLAAL 51  SFGALMIALL DVSSNMAMQP FKMMVGDMVN EEQKXYAYGI QSFLANTGAV 101  VAAILPFVFA YIGLANTAXK GVVPQTVVVA FYVGAALLVI TSAFTIFKVK 151  EYXPETYARY HGIDVAANQE KANWIALLKX A..

Further work revealed the complete nucleotide sequence <SEQ ID 149>:

1 ATGTCGGAAT ATACGCCTCA AACAGCAAAA CAAGGTTTGC CCGCGCTGGC 51 AAAAAGCACGATTTGGATGC TCAGTTTCGG CTTTCTCGGC GTTCAGACGG 101 CCTTTACCCT GCAAAGCTCGCAAATGAGCC GCATTTTTCA AACGCTAGGC 151 GCAGACCCGC ACAATTTGGG CTGGTTTTTCATCCTGCCGC CGCTGGCGGG 201 GATGCTGGTG CAGCCGATTG TCGGCCATTA CTCCGACCGCACTTGGAAGC 251 CGCGTTTGGG CGGCCGCCGT CTGCCGTATC TGCTTTATGG CACGCTGATT301 GCGGTTATTG TGATGATTTT GATGCCGAAC TCGGGCAGCT TCGGTTTCGG 351CTATGCGTCG CTGGCGGCTT TGTCGTTCGG CGCGCTGATG ATTGCGCTGT 401 TAGACGTGTCGTCAAATATG GCGATGCAGC CGTTTAAGAT GATGGTCGGC 451 GACATGGTCA ACGAGGAGCAGAAAGGCTAC GCCTACGGGA TTCAAAGTTT 501 CTTAGCAAAT ACGGGCGCGG TCGTGGCGGCGATTCTGCCG TTTGTGTTTG 551 CGTATATCGG TTTGGCGAAC ACCGCCGAGA AAGGCGTTGTGCCGCAGACC 601 GTGGTCGTGG CGTTTTATGT GGGTGCGGCG TTGCTGGTGA TTACCAGCGC651 GTTCACGATT TTCAAAGTGA AGGAATACGA TCCGGAAACC TACGCCCGTT 701ACCACGGCAT CGATGTCGCC GCGAATCAGG AAAAAGCCAA CTGGATCGAA 751 CTCTTGAAAACCGCGCCTAA GGCGTTTTGG ACGGTTACTT TGGTGCAATT 801 CTTCTGCTGG TTCGCCTTCCAATATATGTG GACTTACTCG GCAGGCGCGA 851 TTGCGGAAAA CGTCTGGCAC ACCACCGATGCGTCTTCCGT AGGTTATCAG 901 GAGGCGGGTA ACTGGTACGG CGTTTTGGCG GCGGTGCAGTCGGTTGCGGC 951 GGTGATTTGT TCGTTTGTAT TGGCGAAAGT GCCGAATAAA TACCATAAGG1001 CGGGTTATTT CGGCTGTTTG GCTTTGGGCG CGCTCGGCTT TTTCTCCGTT 1051TTCTTCATCG GCAACCAATA CGCGCTGGTG TTGTCTTATA CCTTAATCGG 1101 CATCGCTTGGGCGGGCATTA TCACTTATCC GCTGACGATT GTGACCAACG 1151 CCTTGTCGGG CAAGCATATGGGCACTTACT TGGGCTTGTT TAACGGCTCT 1201 ATCTGTATGC CTCAAATCGT CGCTTCGCTGTTGAGTTTCG TGCTTTTCCC 1251 TATGCTGGGC GGCTTGCAGG CCACTATGTT CTTGGTAGGGGGCGTCGTCC 1301 TGCTGCTGGG CGCGTTTTCC GTGTTCCTGA TTAAAGAAAC ACACGGCGGG1351 GTTTGA

This corresponds to the amino acid sequence <SEQ ID 150; ORF16-1>:

1 MSEYTPQTAK QGLPALAKST IWMLSFGFLG VQTAFTLQSS QMSRIFQTLG 51ADPHNLGWFF ILPPLAGMLV QPIVGHYSDR TWKPRLGGRR LPYLLYGTLI 101 AVIVMILMPNSGSFGFGYAS LAALSFGALM IALLDVSSNM AMQPFKMMVG 151 DMVNEEQKGY AYGIQSFLANTGAVVAAILP FVFAYIGLAN TAEKGVVPQT 201 VVVAFYVGAA LLVITSAFTI FKVKEYDPETYARYHGIDVA ANQEKANWIE 251 LLKTAPKAFW TVTLVQFFCW FAFQYMWTYS AGAIAENVWHTTDASSVGYQ 301 EAGNWYGVLA AVQSVAAVIC SFVLAKVPNK YHKAGYFGCL ALGALGFFSV351 FFIGNQYALV LSYTLIGIAW AGIITYPLTI VTNALSGKHM GTYLGLFNGS 401ICMPQIVASL LSFVLFPMLG GLQATMFLVG GVVLLLGAFS VFLIKETHGG 451 V*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. Meningitidis (Strain A)

ORF16 shows 96.7% identity over a 181 as overlap with an ORF (ORF16a)from strain A of N. meningitidis:

The complete length ORF16a nucleotide sequence <SEQ ID 151> is:

1 ATGTCGGAAT ATACGCCTCA AACAGCAAAA CAAGGTTTGC CCGCGCTGGC 51 AAAAAGCACGATTTGGATGC TCAGTTTCGG CTTTCTCGGC GTTCAGACGG 101 CCTTTACCCT GCAAAGCTCGCAGATGAGCC GCATCTTCCA GACGCTCGGT 151 GCCGATCCGC ACAGCCTCGG CTGGTTCTTTATCCTGCCGC CGCTGGCGGG 201 GATGCTGGTG CAGCCGATTG TCGGCCATTA CTCCGACCGCACTTGGAAGC 251 CGCGTTTGGG CGGCCGCCGT CTGCCGTATC TGCTTTATGG CACGCTGATT301 GCGGTTATTG TGATGATTTT GATGCCGAAC TCGGGCAGCT TCGGTTTCGG 351CTATGCGTCG CTGGCGGCTT TGTCGTTCGG CGCGCTGATG ATTGCGCTGT 401 TAGACGTGTCGTCAAATATG GCGATGCAGC CGTTTAAGAT GATGGTCGGC 451 GACATGGTCA ACGAGGAGCAGAAAGGCTAC GCCTACGGGA TTCAAAGTTT 501 CTTAGCGAAT ACGGGCGCGG TCGTGGCGGCGATTCTGCCG TTTGTGTTTG 551 CGTATATCGG TTTGGCGAAC ACCGCCGAGA AAGGCGTTGTGCCGCAGACC 601 GTGGTCGTGG CGTTTTATGT GGGTGCGGCG TTGCTGGTGA TTACCAGCGC651 GTTCACGATT TTCAAAGTGA AGGAATACAA TCCGGAAACC TACGCCCGTT 701ACCACGGCAT CGATGTCGCC GCGAATCAGG AAAAAGCCAA CTGGATCGAA 751 CTCTTGAAAACCGCGCCTAA GGCGTTTTGG ACGGTTACTT TGGTGCAATT 801 CTTCTGCTGG TTCGCCTTCCAATATATGTG GACTTACTCG GCAGGCGCGA 851 TTGCGGAAAA CGTCTGGCAC ACCACCGATGCGTCTTCCGT AGGTTATCAG 901 GAGGCGGGTA ACTGGTACGG CGTTTTGGCG GCGGTGCAGTCGGTTGCGGC 951 GGTGATTTGT TCGTTTGTAT TGGCGAAAGT GCCGAATAAA TACCATAAGG1001 CGGGTTATTT CGGCTGTTTG GCTTTGGGCG CGCTCGGCTT TTTCTCCGTT 1051TTCTTCATCG GCAACCAATA CGCGCTGGTG TTGTCTTATA CCTTAATCGG 1101 CATCGCTTGGGCGGGCATTA TCACTTATCC GCTGACGATT GTGACCAACG 1151 CCTTGTCGGG CAAGCATATGGGCACTTACT TGGGCCTGTT TAACGGCTCT 1201 ATCTGTATGC CGCAAATCGT CGCTTCGCTGTTGAGTTTCG TGCTTTTCCC 1251 TATGCTGGGC GGCTTGCAGG CCACTATGTT CTTGGTAGGGGGCGTCGTCC 1301 TGCTGCTGGG CGCGTTTTCC GTGTTCCTGA TTAAAGAAAC ACACGGCGGG1351 GTTTGA

This encodes a protein having amino acid sequence <SEQ ID 152>:

1 MSEYTPQTAK QGLPALAKST IWMLSFGFLG VQTAFTLQSS QMSRIFQTLG 51ADPHSLGWFF ILPPLAGMLV QPIVGHYSDR TWKPRLGGRR LPYLLYGTLI 101 AVIVMILMPNSGSFGFGYAS LAALSFGALM IALLDVSSNM AMQPFKMMVG 151 DMVNEEQKGY AYGIQSFLANTGAVVAAILP FVFAYIGLAN TAEKGVVPQT 201 VVVAFYVGAA LLVITSAFTI FKVKEYNPETYARYHGIDVA ANQEKANWIE 251 LLKTAPKAFW TVTLVQFFCW FAFQYMWTYS AGAIAENVWHTTDASSVGYQ 301 EAGNWYGVLA AVQSVAAVIC SFVLAKVPNK YHKAGYFGCL ALGALGFFSV351 FFIGNQYALV LSYTLIGIAW AGIITYPLTI VTNALSGKHM GTYLGLFNGS 401ICMPQIVASL LSFVLFPMLG GLQATMFLVG GVVLLLGAFS VFLIKETHGG 451 V*

ORF16a and ORF16-1 show 99.6% identity in 451 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF16 shows 93.9% identity over a 181aa overlap with a predicted ORF(ORF16.ng) from N. gonorrhoeae:

The complete length ORF16ng nucleotide sequence <SEQ ID 153> is:

1 ATGATAGGGG ATCGCCGCGC CGGCAACCAT TTCGGATTTT CCAAAGCAAA 51 TACTTTTCAAATCAAAAAAA AGGATTTACT TTATGTCGGA ATATACGCCT 101 CAAACAGCAA AACAAGGTTTGCCCGCGCCG GCAAAAAGCA CGATTTGGAT 151 GTTGAGCTTC GGCTATCTCG GCGTTCAGACGGCCTTTACC CTGCAAAGCT 201 CGCAGATGAG CCGCATTTTT CAAACGCTAG GCGCAGACCCGCACAATTTG 251 GGCTGGTTTT TCATCCTGCC GCCGCTGGCG GGGATGCTGG TTCAGCCGAT301 AGTGGCTACT ACTCAGACCG CACTTGGAAG CCGCGCTTGG GCGGCCGCCG 351CCTGCCGTAT CTGCTTTACG GCACGCTGAT TGCGGTCATC GTGATGATTT 401 TGATGCCGAACTCGGGCAGC TTCGGTTTCG GCTATGCGTC GCTGGCGGCC 451 TTGTCGTTCG GCGCGCTGATGATTGCGCTG TTGGACGTGT CGTCGAATAT 501 GGCGATGCAG CCGTTTAAGA TGATGGTCGGCGATATGGTC AACGAGGAGC 551 AGAAAAGCTA CGCCTACGGG ATTCAAAGTT TCTTAGCGAATACGGACGCG 601 GTTGTGGCAG CGATTCTGCC GTTTGTGTTC GCGTATATCG GTTTGGCGAA651 CACTGCCGAG AAAGGCGTTG TGCCACAAAC CGTGGTCGTA GCATTCTATG 701TGGGTGCGGC GTTACTGATT ATTACCAGTG CGTTCACAAT CTCCAAAGTC 751 AAAGAATACGACCCGGAAAC CTACGCCCGT TACCACGGCA TCGATGTCGC 801 CGCGAATCAG GAAAAAGCCAACTGGTTCGA ACTCTTAAAA ACCGCGCCTA 851 AAGTGTTTTG GACGGTTACT CCGGTACAGTTTTTCTGCTG GTTCGCCTTC 901 CGGTATATGT GGACTTACTC GGCAGGCGCG ATTGCAGAAAACGTCTGGCA 951 CACTACCGAT GCGTCTTCCG TAGGCCATCA GGAGGCGGGC AACCGGTACG1001 GCGTTTTGGC GGCGGTGTAG

This encodes a protein having amino acid sequence <SEQ ID 154>:

1 MIGDRRAGNH FGFSKANTFQ IKKKDLLYVG IYASNSKTRF ARAGKKHDLD 51 VELRLSRRSDGLYPAKLADE PHFSNARRRP AQFGLVFHPA AAGGDAGSAD 101 SGYYSDRTWKPRLGGRRLPY LLYGTLIAVI VMILMPNSGS FGFGYASLAA 151 LSFGALMIAL LDVSSNMAMQPFKMMVGDMV NEEQKSYAYG IQSFLANTDA 201 VVAAILPFVF AYIGLANTAEKGVVPQTVVV AFYVGAALLI ITSAFTISKV 251 KEYDPETYAR YHGIDVAANQ EKANWFELLKTAPKVFWTVT PVQFFCWFAF 301 RYMWTYSAGA IAENVWHTTD ASSVGHQEAG NRYGVLAAV*

ORF16ng and ORF16-1 show 89.3% identity in 261 aa overlap:

Based on this analysis, including the presence of several putativetransmembrane domains in the gonococcal protein, it is predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 19

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 155>:

1 ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGCATA CCTTGATGCT 51 GAACGGCTGTACGTTGATGT TGTGGGGAAT GAACAACCCG GTCAGCGAAA 101 CAATCACCCG NAAACACGTTGNCAAAGACC AAATCCGNGN CTTCGGTGTG 151 GTTGCCGAAG ACAATGCCCA ATTGGAAAAGGGCAGCCTGG TGATGATGGG 201 CGGAAAATAC TGGTTCGTCG TCAATCCCGA AGATTCGGCGAA.NTGACGG 251 GNATTTTGAN GGCAGGGCTG GACAAACCCT TCCAAATAGT TNAGGATACC301 CCGAGCTATG C.TGCCACCA AGCCCTGCCG GTCAAACTCG GATCGNCTGG 351CAGCCAGAAT...

This corresponds to the amino acid sequence <SEQ ID 156; ORF28>:

1 MLFRKTTAAV LAHTLMLNGC TLMLWGMNNP VSETITRKHV XKDQIRXFGV 51 VAEDNAQLEKGSLVMMGGKY WFVVNPEDSA XXTGILXAGL DKPFQIVXDT 101 PSYXCHQALP VKLGSXGSQN...

Further work revealed the complete nucleotide sequence <SEQ ID 157>:

1 ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA CCTTGATGCT 51 GAACGGCTGTACGTTGATGT TGTGGGGAAT GAACAACCCG GTCAGCGAAA 101 CAATCACCCG CAAACACGTTGACAAAGACC AAATCCGCGC CTTCGGTGTG 151 GTTGCCGAAG ACAATGCCCA ATTGGAAAAGGGCAGCCTGG TGATGATGGG 201 CGGAAAATAC TGGTTCGTCG TCAATCCCGA AGATTCGGCGAAGCTGACGG 251 GCATTTTGAA GGCAGGGCTG GACAAACCCT TCCAAATAGT TGAGGATACC301 CCGAGCTATG CTCGCCACCA AGCCCTGCCG GTCAAACTCG AATCGCCTGG 351CAGCCAGAAT TTCAGTACCG AAGGCCTTTG CCTGCGCTAC GATACCGACA 401 AGCCTGCCGACATCGCCAAG CTGAAACAGC TCGGGTTTGA AGCGGTCAAA 451 CTCGACAATC GGACCATTTACACGCGCTGC GTATCCGCCA AAGGCAAATA 501 CTACGCCACA CCGCAAAAAC TGAACGCCGATTACCATTTT GAGCAAAGTG 551 TGCCTGCCGA TATTTATTAC ACGGTTACTG AAGAACATACCGACAAATCC 601 AAGCTGTTTG CAAATATCTT ATATACGCCC CCCTTTTTGA TACTGGATGC651 GGCGGGCGCG GTACTGGCCT TGCCTGCGGC GGCTCTGGGT GCGGTCGTGG 701ATGCCGCCCG CAAATGA

This corresponds to the amino acid sequence <SEQ ID 158; ORF28-1>:

1 MLFRKTTAAV LAATLMLNGC TLMLWGMNNP VSETITRKHV DKDQIRAFGV 51 VAEDNAQLEKGSLVMMGGKY WFVVNPEDSA KLTGILKAGL DKPFQIVEDT 101 PSYARHQALP VKLESPGSQNFSTEGLCLRY DTDKPADIAK LKQLGFEAVK 151 LDNRTIYTRC VSAKGKYYAT PQKLNADYHFEQSVPADIYY TVTEEHTDKS 201 KLFANILYTP PFLILDAAGA VLALPAAALG AVVDAARK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF28 shows 79.2% identity over a 120aa overlap with an ORF (ORF28a)from strain A of N. meningitidis:

The complete length ORF28a nucleotide sequence <SEQ ID 159> is:

1 ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA CCTTGATGTT 51 GAACGGCTGTACGGTAATGA TGTGGGGTAT GAACAGCCCG TTCAGCGAAA 101 CGACCGCCCG CAAACACGTTGACAAGGACC AAATCCGCGC CTTCGGTGTG 151 GTTGCCGAAG ACAATGCCCA ATTGGAAAAGGGCAGCCTGG TGATGATGGG 201 CGGGAAATAC TGGTTCGTCG TCAATCCTGA AGATTCGGCGAAGCTGACGG 251 GCATTTTGAA GGCCGGGTTG GACAAGCAGT TTCAAATGGT TGAGCCCAAC301 CCGCGCTTTG CCTACCAAGC CCTGCCGGTC AAACTCGAAT CGCCCGCCAG 351CCAGAATTTC AGTACCGAAG GCCTTTGCCT GCGCTACGAT ACCGACAGAC 401 CTGCCGACATCGCCAAGCTG AAACAGCTTG AGTTTGAAGC GGTCGAACTC 451 GACAATCGGA CCATTTACACGCGCTGCGTC TCCGCCAAAG GCAAATACTA 501 CGCCACACCG CAAAAACTGA ACGCCGATTATCATTTTGAG CAAAGTGTGC 551 CTGCCGATAT TTATTACACG GTTACGAAAA AACATACCGACAAATCCAAG 601 TTGTTTGAAA ATATTGCATA TACGCCCACC ACGTTGATAC TGGATGCGGT651 GGGCGCGGTG CTGGCCTTGC CTGTCGCGGC GTTGATTGCA GCCACGAATT 701CCTCAGACAA ATGA

This encodes a protein having amino acid sequence <SEQ ID 160>:

1 MLFRKTTAAV LAATLMLNGC TVMMWGMNSP FSETTARKHV DKDQIRAFGV 51 VAEDNAQLEKGSLVMMGGKY WFVVNPEDSA KLTGILKAGL DKQFQMVEPN 101 PRFAYQALPV KLESPASQNFSTEGLCLRYD TDRPADIAKL KQLEFEAVEL 151 DNRTIYTRCV SAKGKYYATP QKLNADYHFEQSVPADIYYT VTKKHTDKSK 201 LFENIAYTPT TLILDAVGAV LALPVAALIA ATNSSDK*

ORF28a and ORF28-1 show 86.1% identity in 238 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF28 shows 84.2% identity over a 120aa overlap with a predicted ORF(ORF28.ng) from N. gonorrhoeae:

The complete length ORF28ng nucleotide sequence <SEQ ID 161> is

1 ATGTTGTTCC GTAAAACGAC CGCCGCCGTT TTGGCGGCAA CCTTGATACT 51 GAACGGCTGTACGATGATGT TGCGGGGGAT GAACAACCCG GTCAGCCAAA 101 CAATCACCCG CAAACACGTTGACAAAGACC AAATCCGCGC CTTCGGTGTG 151 GTTGCCGAAG ACAATGCCCA ATTGGAAAAGGGCAGCCTGG TGATGATGGG 201 CGGGAAATAC TGGTTCGCCG TCAATCCCGA AGATTCGGCGAAGCTGACGG 251 GCCTTTTGAA GGCCGGGTTG GACAAGCCCT TCCAAATAGT TGAGGATACC301 CCGAGCTATG CCCGCCACCA AGCCCTGCCG GTCAAATTCG AAGCGCCCGG 351CAGCCAGAAT TTCAGTACCG GAGGTCTTTG CCTGCGCTAT GATACCGGCA 401 GACCTGACGACATCGCCAAG CTGAAACAGC TTGAGTTTAA AGCGGTCAAA 451 CTCGACAATC GGACCATTTACACGCGCTGC GTATCCGCCA AAGGCAAATA 501 CTACGCCACG CCGCAAAAAC TGAACGCCGATTATCATTTT GAGCAAAGTG 551 TGCCCGCCGA TATTTATTAT ACGGTTACTG AAAAACATACCGACAAATCC 601 AAGCTGTTTG GAAATATCTT ATATACGCCC CCCTTGTTGA TATTGGATGC651 GGCGGCCGCG GTGCTGGTCT TGCCTATGGC TCTGATTGCA GCCGCGAATT 701CCTCAGACAA ATGA

This encodes a protein having amino acid sequence <SEQ ID 162>:

1 MLFRKTTAAV LAATLILNGC TMMLRGMNNP VSQTITRKHV DKDQIRAFGV 51 VAEDNAQLEKGSLVMMGGKY WFAVNPEDSA KLTGLLKAGL DKPFQIVEDT 101 PSYARHQALP VKFEAPGSQNFSTGGLCLRY DTGRPDDIAK LKQLEFKAVK 151 LDNRTIYTRC VSAKGKYYAT PQKLNADYHFEQSVPADIYY TVTEKHTDKS 201 KLFGNILYTP PLLILDAAAA VLVLPMALIA AANSSDK*

ORF28ng and ORF28-1 share 90.0% identity in 231 aa overlap:

Based on this analysis, including the presence of a putativetransmembrane domain in the gonococcal protein, it was predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

ORF28-1 (24 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 6A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 6B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for ELISA, whichgave a positive result. These experiments confirm that ORF28-1 is asurface-exposed protein, and that it may be a useful immunogen.

Example 20

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 163>:

1 ..GTCAGTCCTG TACTGCCTAT TACACACGAA CGGACAGGGT TTGAAGGTGT 51  TATCGGTTAT GAAACCCATT TTTCAGGGCA CGGACATGAA GTACACAGTC 101  CGTTCGATCA TCATGATTCA AAAAGCACTT CTGATTTCAG CGGCGGTGTA 151  GACGGCGGTT TTACTGTTTA CCAACTTCAT CGAACATGGT CGGAAATCCA 201  TCCGGAGGAT GAATATGACG GGCCGCAAGC AGCG.ATTAT CCGCCCCCCG 251  GAGGAGCAAG GGATATATAC AGCTATTATG TCAAAGGAAC TTCAACAAAA 301  ACAAAGACTA GTATTGTCCC TCAAGCCCCA TTTTCAGACC GTTGGCTAGA 351  AGAAAATGCC GGTGCCGCCT CTGGT..

This corresponds to the amino acid sequence <SEQ ID 164; ORF29>:

1 ..VSPVLPITHE RTGFEGVIGY ETHFSGHGHE VHSPFDHHDS KSTSDFSGGV 51  DGGFTVYQLH RTWSEIHPED EYDGPQAAXY PPPGGARDIY SYYVKGTSTK 101  TKTSIVPQAP FSDRWLEENA GAASG..

Further work revealed the complete nucleotide sequence <SEQ ID 165>:

1 ATGAATTTGC CTATTCAAAA ATTCATGATG CTGTTTGCAG CAGCAATATC 51 GTTGCTGCAAATCCCCATTA GTCATGCGAA CGGTTTGGAT GCCCGTTTGC 101 GCGATGATAT GCAGGCAAAACACTACGAAC CGGGTGGTAA ATACCATCTG 151 TTTGGTAATG CTCGCGGCAG TGTTAAAAAGCGGGTTTACG CCGTCCAGAC 201 ATTTGATGCA ACTGCGGTCA GTCCTGTACT GCCTATTACACACGAACGGA 251 CAGGGTTTGA AGGTGTTATC GGTTATGAAA CCCATTTTTC AGGGCACGGA301 CATGAAGTAC ACAGTCCGTT CGATCATCAT GATTCAAAAA GCACTTCTGA 351TTTCAGCGGC GGTGTAGACG GCGGTTTTAC TGTTTACCAA CTTCATCGAA 401 CAGGGTCGGAAATCCATCCG GAGGATGGAT ATGACGGGCC GCAAGGCAGC 451 GATTATCCGC CCCCCGGAGGAGCAAGGGAT ATATACAGCT ATTATGTCAA 501 AGGAACTTCA ACAAAAACAA AGACTAATATTGTCCCTCAA GCCCCATTTT 551 CAGACCGTTG GCTAAAAGAA AATGCCGGTG CCGCCTCTGGTTTTTTCAGC 601 CGTGCGGATG AAGCAGGAAA ACTGATATGG GAAAGCGACC CCAATAAAAA651 TTGGTGGGCT AACCGTATGG ATGATGTTCG CGGCATCGTC CAAGGTGCGG 701TTAATCCTTT TTTAATGGGT TTTCAAGGAG TAGGGATTGG GGCAATTACA 751 GACAGTGCAGTAAGCCCGGT CACAGATACA GCCGCGCAGC AGACTCTACA 801 AGGTATTAAT GATTTAGGAAAATTAAGTCC GGAAGCACAA CTTGCTGCCG 851 CGAGCCTATT ACAGGACAGT GCTTTTGCGGTAAAAGACGG TATCAACTCT 901 GCCAAACAAT GGGCTGATGC CCATCCAAAT ATAACAGCTACTGCCCAAAC 951 TGCCCTTTCC GCAGCAGAGG CCGCAGGTAC GGTTTGGAGA GGTAAAAAAG1001 TAGAACTTAA CCCGACTAAA TGGGATTGGG TTAAAAATAC CGGTTATAAA 1051AAACCTGCTG CCCGCCATAT GCAGACTTTA GATGGGGAGA TGGCAGGTGG 1101 GAATAAACCTATTAAATCTT TACCAAACAG TGCCGCTGAA AAAAGAAAAC 1151 AAAATTTTGA GAAGTTTAATAGTAACTGGA GTTCAGCAAG TTTTGATTCA 1201 GTGCACAAAA CACTAACTCC CAATGCACCTGGTATTTTAA GTCCTGATAA 1251 AGTTAAAACT CGATACACTA GTTTAGATGG AAAAATTACAATTATAAAAG 1301 ATAACGAAAA CAACTATTTT AGAATCCATG ATAATTCACG AAAACAGTAT1351 CTTGATTCAA ATGGTAATGC TGTGAAAACC GGTAATTTAC AAGGTAAGCA 1401AGCAAAAGAT TATTTACAAC AACAAACTCA TATCAGGAAC TTAGACAAAT 1451 GA

This corresponds to the amino acid sequence <SEQ ID 166; ORF29-1>:

1 MNLPIQKFMM LFAAAISLLQ IPISHANGLD ARLRDDMQAK HYEPGGKYHL 51 FGNARGSVKKRVYAVQTFDA TAVSPVLPIT HERTGFEGVI GYETHFSGHG 101 HEVHSPFDHH DSKSTSDFSGGVDGGFTVYQ LHRTGSEIHP EDGYDGPQGS 151 DYPPPGGARD IYSYYVKGTS TKTKTNIVPQAPFSDRWLKE NAGAASGFFS 201 RADEAGKLIW ESDPNKNWWA NRMDDVRGIV QGAVNPFLMGFQGVGIGAIT 251 DSAVSPVTDT AAQQTLQGIN DLGKLSPEAQ LAAASLLQDS AFAVKDGINS301 AKQWADAHPN ITATAQTALS AAEAAGTVWR GKKVELNPTK WDWVKNTGYK 351KPAARHMQTL DGEMAGGNKP IKSLPNSAAE KRKQNFEKFN SNWSSASFDS 401 VHKTLTPNAPGILSPDKVKT RYTSLDGKIT IIKDNENNYF RIHDNSRKQY 451 LDSNGNAVKT GNLQGKQAKDYLQQQTHIRN LDK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF29 shows 88.0% identity over a 125aa overlap with an ORF (ORF29a)from strain A of N. meningitidis:

The complete length ORF29a nucleotide sequence <SEQ ID 167> is:

1 ATGAATTNGC CTATTCAAAA ATTCATGATG CTGTTTGCAG CAGCAATATC 51 GTNGCTGCAAATCCCNATTA GTCATGCGAA CGGTTTGGAT GCCCGTTTGC 101 GCGATGATAT GCAGGCAAAACACTACGAAC CGGGTGGTAA ATACCATCTG 151 TTTGGTAATG CTCGCGGCAG TGTTAAAAATCGGGTTTACG CCGTCCAAAC 201 ATTTGATGCA ACTGCGGTCG GCCCCATACT GCCTATTACACACGAACGGA 251 CAGGATTTGA AGGCATTATC GGTTATGAAA CCCATTTTTC AGGACATGGA301 CATGAAGTAC ACAGTCCGTT CGATAATCAT GATTCAAAAA GCACTTCTGA 351TTTCAGCGGC GGCGTAGACG GTGGTTTTAC CGTTTACCAA CTTCATCGGA 401 CAGGGTCGGAAATCCATCCG GAGGATGGAT ATGACGGGCC GCAAGGCAGC 451 GATTATCCGC CCCCCGGAGGAGCAAGGGAT ATATACANNT ANTATGTCAA 501 AGGAACTTCA ACAAAAACAA AGAGTAATATTGTTCCCCGA GCCCCATTTT 551 CAGACCGCTG GCTAAAAGAA AATGCCGGTG CCGCCTCTGGTTTTTTCAGC 601 CGTGCTGATG AAGCAGGAAA ACTGATATGG GAAAGCGACC CCAATAAAAA651 TTGGTGGGCT AACCGTATGG ATGATATTCG CGGCATCGTC CAAGGTGCGG 701TTAATCCTTT TTTAATGGGT TTTCAAGGAG TAGGGATTGG GGCAATTACA 751 GACAGTGCAGTAAGCCCGGT CACAGATACA GCCGCGCAGC AGACTCTACA 801 AGGTATNAAT CATTTAGGAAANTTAAGTCC CGAAGCACAA CTTGCGGCTG 851 CAACCGCATT ACAAGACAGT GCTTTTGCGGTAAAAGACGG TATCAATTCC 901 GCCAGACAAT GGGCTGATGC CCATCCGAAT ATAACTGCAACAGCCCAAAC 951 TGCCCTTGCC GTAGCAGANG CCGCAACTAC GGTTTGGGGC GGTAAAAAAG1001 TAGAACTTAA CCCGACCAAA TGGGATTGGG TTAAAAATAC NGGCTATAAN 1051ACACCTGCTG TTCGCACCAT GCATACTTTG GATGGGGAAA TGGCCGGTGG 1101 GAATAGACCGCCTAAATCTA TAACGTCCAA CAGCAAAGCA GATGCTTCCA 1151 CACAACCGTC TTTACAAGCGCAACTAATTG GAGAACAAAT TANNNNNGGG 1201 CATGCTTATA ACAAGCATGT CATAAGACAACAAGAATTTA CGGATTTAAA 1251 TATCAATTCA CCAGCAGATT TTGCTCGGCA TATTGAAAATATTGTTAGCC 1301 ATCCANCAAA TATGAAAGAG TTACCTCGCG GTAGAACTGC GTATTGGGAT1351 NATAAAACAG GGACNATAGT TATCCGAGAT AAAAATTCTG ACGATGGAGG 1401TACAGCATTT AGACCAACAT CAGGTAAAAA ATATTATGAT GATTTATAG

This encodes a protein having amino acid sequence <SEQ ID 168>:

1 MNXPIQKFMM LFAAAISXLQ IPISHANGLD ARLRDDMQAK HYEPGGKYHL 51 FGNARGSVKNRVYAVQTFDA TAVGPILPIT HERTGFEGII GYETHFSGHG 101 HEVHSPFDNH DSKSTSDFSGGVDGGFTVYQ LHRTGSEIHP EDGYDGPQGS 151 DYPPPGGARD IYXXYVKGTS TKTKSNIVPRAPFSDRWLKE NAGAASGFFS 201 RADEAGKLIW ESDPNKNWWA NRMDDIRGIV QGAVNPFLMGFQGVGIGAIT 251 DSAVSPVTDT AAQQTLQGXN HLGXLSPEAQ LAAATALQDS AFAVKDGINS301 ARQWADAHPN ITATAQTALA VAXAATTVWG GKKVELNPTK WDWVKNTGYX 351TPAVRTMHTL DGEMAGGNRP PKSITSNSKA DASTQPSLQA QLIGEQIXXG 401 HAYNKHVIRQQEFTDLNINS PADFARHIEN IVSHPXNMKE LPRGRTAYWD 451 XKTGTIVIRD KNSDDGGTAFRPTSGKKYYD DL*

ORF29a and ORF29-1 show 90.1% identity in 385 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF29 shows 88.8% identity over a 125aa overlap with a predicted ORF(ORF29.ng) from N. gonorrhoeae:

The complete length ORF29ng nucleotide sequence <SEQ ID 169> ispredicted to encode a protein having amino acid sequence <SEQ ID 170>:

  1 MNLPIQKFMM LFAAAISLLQ IPISHANGLD ARLRDDMQAK HYEPGGKYHL 51 FGNARGSVKN RVCAVQTFDA TAVGPILPIT HERTGFEGVI GYETHFSGHG101 HEVHSPFDNH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP EDGYDGPQGG151 GYPPPGGARD IYSYHIKGTS TKTKINTVPQ APFSDRWLKE NAGAASGFLS201 RADEAGKLIW ENDPDKNWRA NRMDDIRGIV QGAVNPFLTG FQGLGVGAIT251 DSAVSPVTYA AARKTLQGIH NLGNLSPEAQ LAAATALQDS AFAVKDSINS301 ARQWADAHPN ITATAQTALA VTEAATTVWG GKKVELNPAK WDWVKNTGYK351 KPAARHMQTV DGEMAGGNKP LESKNTVTTN NFFENTGYTE KVLRQASNGD401 YHGFPQSVDA FSENGTVIQI VGGDNIVRHK LYIPGSYKGK DGNFEYIREA451 DGKINHRLFV PNQQLPEK*

In a second experiment, the following DNA sequence <SEQ ID 171> wasidentified:

   1 atgAATTTGC CTATTCAAAA ATTCATGATG ctgttggcAg cggcaatatc  51 gatgctGCat ATCCCCATTA GTCATGCGAA CGGTTTGGAT GCCCGTTTGC 101 GCGATGATAT GCAGGCAAAA CACTACGAAC CGGGTGGCAA ATACCATCTG 151 TTTGGTAATG CTCGCGGCAG TGTTAAAAAT CGGGTTTGCG CCGTCCAAAC 201 ATTTGATGCA ACTGCGGTCG GCCCCATACT GCCTATTACA CACGAACGGA 251 CAGGATTTGA AGGTGTTATC GGCTATGAAA CCCATTTTTC AGGACACGGA 301 CACGAAGTAC ACAGTCCGTT CGATAATCAT GATTCAAAAA GCACTTCTGA 351 TTTCAGCGGC GGCGTAGACG GCGGTTTTAC CGTTTACCAA CTTCATCGGA 401 CAGGGTCGGA AATACATCCC GCAGACGGAT ATGACGGGCC TCAAGGCGGC 451 GGTTATCCGG AACCACAAGG GGCAAGGGAT ATATACAGCT ACCATATCAA 501 AGGAACTTCA ACCAAAACAA AGATAAACAC TGTTCCGCAA GCCCCTTTTT 551 CAGACCGCTG GCTAAAAGAA AATGCCGGTG CCGCTTCCGG TTTTCTCAGC 601 CGTGCGGATG AAGCAGGAAA ACTGATATGG GAAAACGACC CCGATAAAAA 651 TTGGCGGGCT AACCGTATGG ATGATATTCG CGGCATCGTC CAAGGTGCGG 701 TTAATCCTTT TTTAACGGGT TTTCAAGGGG TAGGGATTGG GGCAATTACA 751 GACAGTGCGG TAAGCCCGGT CACAGATACA GCCGCTCAGC AGACTCTACA 801 AGGTATTAAT GATTTAGGAA ATTTAAGTCC GGAAGCACAA CTTGCCGCCG 851 CGAGCCTATT ACAGGACAGT GCCTTTGCGG TAAAAGACGG CATCAATTCC 901 GCCAGACAAT GGGCTGATGC CCATCCGAAT ATAACAGCAA CAGCCCAAAC 951 TGCCCTTGCC GTAGCAGAGG CCGCAGGTAC GGTTTGGCGC GGTAAAAAAG1001 TAGAACTTAA CCCGACCAAA TGGGATTGGG TTAAAAATAC CGGCTATAAA1051 AAACCTGCTG CCCGCCATAT GCAGACTGTA GATGGGGAGA TGGCAGGGGG1101 GAATAGACCG CCTAAATCTA TAACGTCGGA AGGAAAAGCT AATGCTGCAA1151 CCTATCCTAA GTTGGTTAAT CAGCTAAATG AGCAAAACTT AAATAACATT1201 GCGGCTCAAG ATCCAAGATT GAGTCTAGCT ATTCATGAGG GTAAAAAAAA1251 TTTTCCAATA GGAACTGCAA CTTATGAAGA GGCAGATAGA CTAGGTAAAA1301 TTTGGGTTGG TGAGGGTGCA AGACAAACTA GTGGAGGCGG ATGGTTAAGT1351 AGAGATGGCA CTCGACAATA TCGGCCACCA ACAGAAAAAA AATCACAATT1401 TGCAACTACA GGTATTCAAG CAAATTTTGA AACTTATACT ATTGATTCAA1451 ATGAAAAAAG AAATAAAATT AAAAATGGAC ATTTAAATAT TAGGTAA

This encodes a protein having amino acid sequence <SEQ ID 172;ORF29ng-1>:

  1 MNLPIQKFMM LLAAAISMLH IPISHANGLD ARLRDDMQAK HYEPGGKYHL 51 FGNARGSVKN RVCAVQTFDA TAVGPILPIT HERTGFEGVI GYETHFSGHG101 HEVHSPFDNH DSKSTSDFSG GVDGGFTVYQ LHRTGSEIHP ADGYDGPQGG151 GYPEPQGARD IYSYHIKGTS TKTKINTVPQ APFSDRWLKE NAGAASGFLS201 RADEAGKLIW ENDPDKNWRA NRMDDIRGIV QGAVNPFLTG FQGVGIGAIT251 DSAVSPVTDT AAQQTLQGIN DLGNLSPEAQ LAAASLLQDS AFAVKDGINS301 ARQWADAHPN ITATAQTALA VAEAAGTVWR GKKVELNPTK WDWVKNTGYK351 KPAARHMQTV DGEMAGGNRP PKSITSEGKA NAATYPKLVN QLNEQNLNNI401 AAQDPRLSLA IHEGKKNFPI GTATYEEADR LGKIWVGEGA RQTSGGGWLS451 RDGTRQYRPP TEKKSQFATT GIQANFETYT IDSNEKRNKI KNGHLNIR*

ORF29ng-1 and ORF29-1 show 86.0% identity in 401 aa overlap:

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it is predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

Example 21

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 173>:

  1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC 51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAATGTTCC101 ACACGCGGGC AGATGCACCG ATGCAG...

This corresponds to the amino acid sequence <SEQ ID 174; ORF30>:

1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QMFHTRADAP MQ..

Further work revealed the complete nucleotide sequence <SEQ ID 175>:

  1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC 51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC101 ACACGCGGGC AGATGCACCG ATGCAGTTGG CGGAGCTTTC TCAAAAGGAG151 ATGAAGGAGA CAGAGGGGGC GTTTCTTCCA TTGGCTATCT TGGGTGGTGC201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA251 GACCAGCTTC TGTTAGAGAT GTTGCTATTG CTGGCGGATT AGGCGCAATT301 CCTGGTGGTG TAGGCGCCGC AGGAAAGGTT GTTTCCTTTG CTAAATATGG351 ACGTGAGATT AAAATCGGCA ATAATATGCG GATAGCCCCT TTCGGTAATA401 GAACAGGTCA TCCTATTGGA AAATTTCCCC ATTATCATCG TCGAGTTACG451 GATAATACGG GCAAGACTTT GCCTGGACAG GGAATTGGTC GTCATCGCCC501 TTGGGAATCA AAATCTACGG ACAGATCATG GAAAAACCGC TTCTAA

This corresponds to the amino acid sequence <SEQ ID 176; ORF30-1>:

  1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE 51 MKETEGAFLP LAILGGAAIG MWTQHGFSYA TTGRPASVRD VAIAGGLGAI101 PGGVGAAGKV VSFAKYGREI KIGNNMRIAP FGNRTGHPIG KFPHYHRRVT151 DNTGKTLPGQ GIGRHRPWES KSTDRSWKNR F*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF30 shows 97.6% identity over a 42aa overlap with an ORF (ORF30a) fromstrain A of N. meningitidis:

The complete length ORF30a nucleotide sequence <SEQ ID 177> is:

  1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATTGCCCC 51 CGCAATGGCA AACGGCTTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC101 ACACGCGGGC AGATGCACCG ATGCAGTTGG CGGAGCTTTC TCAAAAGGAG151 ATGAAGGANA CAGNGGGGGC GTTTCTTCCA TTGGNTATCT TGGGTGGTGC201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA251 GACCAGCTTC TGTTAGAGAT GTTGCTATTG CTGGCGGATT AGGCGCAATT301 CCTGGTGNTG TAGGCGCCGC AGGAAAGGTT GTTTCCTTTG CTAAATATGG351 ACGTGAGATT AAAATCGGCA ATAATATGCG GATAGCCCCT TTCGGTAATA401 GAACAGGTCA TCCTATTGGN AAATTTCCCC ATTATCATCG TCGAGTTACG451 GATAATACGG GCAAGACTTT GCCTGGACAG GGAATTGGTC GTCATCGCCC501 TTGGGAATCA AAATCTACGG ACAGATCATG GAAAAACCGC TTCTAA

This encodes a protein having amino acid sequence <SEQ ID 178>:

  1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE 51 MKXTXGAFLP LXILGGAAIG MWTQHGFSYA TTGRPASVRD VAIAGGLGAI101 PGXVGAAGKV VSFAKYGREI KIGNNMRIAP FGNRTGHPIG KFPHYHRRVT151 DNTGKTLPGQ GIGRHRPWES KSTDRSWKNR F*

ORF30a and ORF30-1 show 97.8% identity in 181 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF30 shows 97.6% identity over a 42aa overlap with a predicted ORF(ORF30.ng) from N. gonorrhoeae:

The complete length ORF30ng nucleotide sequence <SEQ ID 179> is

  1 ATGAAAAAAC AAATCACCGC AGCCGTAATG ATGCTGTCTA TGATCGCCCC 51 CGCAATGGCA AACGGATTGG ACAATCAGGC ATTTGAAGAC CAAGTGTTCC101 ACACGCGGGC AGATGCGCCG ATGCAGTTGG CGGAGCTTTC TCAGAAGGAG151 ATGAAGGAGA CTGAAGGGGC TTTTCTTCCA TTGGCTATCT TGGGTGGTGC201 TGCCATTGGT ATGTGGACAC AGCATGGTTT TAGTTATGCA ACGACAGGCA251 GACCAGCTTC TGTTAGAGAT GTTGCTGGCG GATTAGGCGC AATTCCTGGT301 GATGTAGGTG CTGCAGGAAA GGTTGTTTCC TTTGCTAAAT ATGGACGTGA351 GATTAAAATC GGCAATAATA TGCGGATAGC CCCTTTCGGT AATAGAACAG401 GTCATCCTAT TGGAAAATTT CCCCATTATC ATCGTCGAGT TACGGATAAT451 ACGGGCAAGA CTTTGCCTGG ACAGGGAATT GGTCGTCATC GCCCTTGGGA501 ATCAAAATCT ACGGACAGAT CATGGAAAAA CCGCTTCTAA

This encodes a protein having amino acid sequence <SEQ ID 180>:

  1 MKKQITAAVM MLSMIAPAMA NGLDNQAFED QVFHTRADAP MQLAELSQKE 51 MKETEGAFLP LAILGGAAIG MWTQHGFSYA TTGRPASVRD VAGGLGAIPG101 DVGAAGKVVS FAKYGREIKI GNNMRIAPFG NRTGHPIGKF PHYHRRVTDN151 TGKTLPGQGI GRHRPWESKS TDRSWKNRF*

ORF30ng and ORF30-1 show 98.3% identity in 181 aa overlap:

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it is predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

Example 22

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 181>:

  1 ATGAATAAAA CTCTCTATCG TGTAATTTTC AACCGCAAAC GTGGGGCTGT 51 GrTAGCCGTT GCTGAAACTA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA101 GTGATTCAGG CAGCGCTCAT GTGAAATCTG TTCCTTTTGG TACTACTCAT151 GCACCTGTTT GTg.CGTTaC AAATATCTTT TCTTTTTCTT TATTGGGCTT201 TTCTTTATGT TTGGCTGTAG GtacGGyCAA TATTGCTTTT GCTGATGGCA 251 TT..

This corresponds to the amino acid sequence <SEQ ID 182; ORF31>:

 1 MNKTLYRVIF NRKRGAVXAV AETTKREGKS CADSDSGSAH VKSVPFGTTH51 APVCXVTNIF SFSLLGFSLC LAVGTXNIAF ADGI..

Further work revealed a further partial nucleotide sequence <SEQ ID183>:

  1 ATGAATAAAA CTCTCTATCG TGTAATTTTC AACCGCAAAC GTGGGGCTGT 51 GGTAGCCGTT GCTGAAACTA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA101 GTGATTCAGG CAGCGCTCAT GTGAAATCTG TTCCTTTTGG TACTACTCAT151 GCACCTGTTT GTCGTTCAAA TATCTTTTCT TTTTCTTTAT TGGGCTTTTC201 TTTATGTTTG GCTGTAGGTA CGGCCAATAT TGCTTTTGCT GATGGCATT..

This corresponds to the amino acid sequence <SEQ ID 184; ORF31-1>:

-   -   1 MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSDSGSAH VKSVPFGTTH    -   51 APVCRSNIFS FSLLGFSLCL AVGTANIAFA DGI . . . .

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. gonorrhoeae

ORF31 shows 76.2% identity over a 84aa overlap with a predicted ORF(ORF31.ng) from N. gonorrhoeae:

The complete length ORF31ng nucleotide sequence <SEQ ID 185> is:

  1 ATGAACAAAA CCCTCTATCG TGTGATTTTC AACCGCAAAC GCGGTGCTGT 51 GGTAGCTGTT GCCGAAACCA CCAAGCGCGA AGGTAAAAGC TGTGCCGATA101 GTGGTTCGGG CAGCGTTTAT GTGAAATCCG TTTCTTTCAT TCCTACTCAT151 TCCAAAGCCT TTTGTTTTTC TGCATTAGGC TTTTCTTTAT GTTTGGCTTT201 GGGTACGGTC AATATTGCTT TTGCTGACGG CATTATTACT GATAAAGCTG251 CTCCTAAAAC CCAACAAGCC ACGATTCTGC AAACAGGTaa cGGCATACCG301 CAAGTCAATA TTCAAACCCC TACTTCGGCA GGGGTTTCTG TTAATCAATA351 TGCCCAGTTT GATGTGGGTA ATCGCGGGGC GATTTTAAAC AACAGTCGCA401 GCAACACCCA AACACAGCTA GGCGGTTGGA TTCAAGGCAA TCCTTGGTTG451 ACAAGGGGCG AAGCACGTGT GGTTGTAAAC CAAATCAACA GCAGCCATCC501 TTCACAACTG AATGGCTATA TTGAAGTGGG TGGACGACGT GCAGAAGTCG551 TTATTGCCAA TCCGGCAGGG ATTGCAGTCA ATGGTGGTGG TTTTATCAAT601 GCTTCCCGTG CCACTTTGAC GACAGGCCAA CCGCAATATC AAGCAGGAGA651 CTTTAGCGGC TTTAAGATAA GGCAAGGCAA TGCTGTAATC GCCGGACACG701 GTTTGGATGC CCGTGATACC GATTTCACAC GTATTCTTGT ATGCCAACAA751 AATCACCTTG ATCAGTACGG CCGAACAAGC AGGCATTCGT AA

This encodes a protein having amino acid sequence <SEQ ID 186>:

1 MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSGSGSVY VKSVSFIPTH 51 SKAFCFSALGFSLCLALGTV NIAFADGIIT DKAAPKTQQA TILQTGNGIP 101 QVNIQTPTSA GVSVNQYAQFDVGNRGAILN NSRSNTQTQL GGWIQGNPWL 151 TRGEARVVVN QINSSHPSQL NGYIEVGGRRAEVVIANPAG IAVNGGGFIN 201 ASRATLTTGQ PQYQAGDFSG FKIRQGNAVI AGHGLDARDTDFTRILVCQQ 251 NHLDQYGRTS RHS*

This gonococcal protein shares 50% identity over a 149aa overlap withthe pore-forming hemolysins-like HecA protein from Erwinia chrysanthemi(accession number L39897):

orf31ng 96 GNGIPQVNIQTPTSAGVSVNQYAQFDVGNRGAILNNSRSN-TQTQLGGWIQGNPWLTRGE154 GNG+P VNI TP ++G+S N+Y  F+V NRG ILNN  +  T +QLGG IQ NP L HecA 45GNGVPVVNIATPDASGLSHNRYHDFNVDNRGLILNNGTARLTPSQLGGLIQNNPNLNGRA 104 Orf31ng155 ARVVVNQINSSHPSQLNGYIEVGGRRAEVVIANPAGIAVNGGGFINASRATLTTGQPQYQ 214A  ++N++ S + S+L GY+EV G+ A VV+ANP GI  +G GF+N  R TLTTG PQ+ HecA 105AAAILNEVVSPNRSRLAGYLEVAGQAANVVVANPYGITCSGCGFLNTPRLTLTTGTPQFD 164 Orf31ng215 -AGDFSGFKIRQGNAVIAGHGLDARDTDF 242  AG  SG  +R G+ +I G GLDA  +D+ HecA165 AAGGLSGLDVRGGDILIDGAGLDASRSDY 193

Furthermore, ORF31ng and ORF31-1 show 79.5% identity in 83 aa overlap:

On this basis, including the homology with hemolysins, and also withadhesins, it is predicted that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 23

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 187>:

1 ATGAATACTC CTCCTTTTGT CTGTTGGATT TTTTGCAAGG TCATCGACAA 51 TTTCGGCGACATCGGCGTTT CGTGGCGGCT CGCCCGTGTT TTGCACCGCG 101 AACTCGGTTG GCAGGTGCATTTGTGGACGG ACGATGTGTC CGCCTTGCGT 151 GCGCTTTGCC CTGATTTGCC CGATGTTCCCTGCGTTCATC AGGATATTCA 201 TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACCGCG..

This corresponds to the amino acid sequence <SEQ ID 188; ORF32>:

1 MNTPPFVCWI FCKVIDNFGD IGVSWRLARV LHRELGWQVH LWTDDVSALR 51 ALCPDLPDVPCVHQDIHVRT WHSDAADIDT A..

Further work revealed the complete nucleotide sequence <SEQ ID 189>:

1 ATGAATACTC CTCCTTTTGT CTGTTGGATT TTTTGCAAGG TCATCGACAA 51 TTTCGGCGACATCGGCGTTT CGTGGCGGCT CGCCCGTGTT TTGCACCGCG 101 AACTCGGTTG GCAGGTGCATTTGTGGACGG ACGATGTGTC CGCCTTGCGT 151 GCGCTTTGCC CTGATTTGCC CGATGTTCCCTGCGTTCATC AGGATATTCA 201 TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACCGCGCCTGTTC 251 CCGATGTCGT CATCGAAACT TTTGCCTGCG ACCTGCCCGA AAATGTGCTG301 CACATTATCC GCCGACACAA GCCGCTTTGG CTGAATTGGG AATATTTGAG 351CGCGGAGGAA AGCAATGAAA GGCTGCATCT GATGCCTTCG CCGCAGGAGG 401 GTGTTCAAAAATATTTTTGG TTTATGGGTT TCAGCGAAAA AAGCGGCGGG 451 TTGATACGCG AACGTGATTACTGCGAAGCC GTCCGTTTCG ATACTGAAGC 501 CCTGCGAGAG CGGCTGATGC TGCCCGAAAAAAACGCCTCC GAATGGCTGC 551 TTTTCGGCTA TCGGAGCGAT GTTTGGGCAA AGTGGCTGGAAATGTGGCGA 601 CAGGCAGGCA GCCCGATGAC ACTGTTGCTG GCGGGGACGC AAATCATCGA651 CAGCCTCAAA CAAAGCGGCG TTATTCCGCA AGATGCCCTG CAAAACGACG 701GCGATGTTTT TCAGACGGCA TCCGTCCGCC TCGTCAAAAT CCCTTTCGTG 751 CCGCAACAGGACTTCGACCA ACTGCTGCAC CTTGCCGACT GCGCCGTCAT 801 CCGCGGCGAA GACAGTTTCGTGCGCGCCCA GCTTGCGGGC AAACCCTTCT 851 TTTGGCACAT CTACCCGCAA GACGAGAATGTCCATCTCGA CAAACTCCAC 901 GCCTTTTGGG ATAAGGCACA CGGTTTCTAC ACGCCCGAAACCGTGTCGGC 951 ACACCGCCGT CTTTCGGACG ACCTCAACGG CGGAGAGGCT TTATCCGCAA1001 CACAACGCCT CGAATGTTGG CAAACCCTGC AACAACATCA AAACGGCTGG 1051CGGCAAGGCG CGGAGGATTG GAGCCGTTAT CTTTTCGGGC AGCCGTCAGC 1101 TCCTGAAAAACTCGCTGCCT TTGTTTCAAA GCATCAAAAA ATACGCTAG

This corresponds to the amino acid sequence <SEQ ID 190; ORF32-1>:

1 MNTPPFVCWI FCKVIDNFGD IGVSWRLARV LHRELGWQVH LWTDDVSALR 51 ALCPDLPDVPCVHQDIHVRT WHSDAADIDT APVPDVVIET FACDLPENVL 101 HIIRRHKPLW LNWEYLSAEESNERLHLMPS PQEGVQKYFW FMGFSEKSGG 151 LIRERDYCEA VRFDTEALRE RLMLPEKNASEWLLFGYRSD VWAKWLEMWR 201 QAGSPMTLLL AGTQIIDSLK QSGVIPQDAL QNDGDVFQTASVRLVKIPFV 251 PQQDFDQLLH LADCAVIRGE DSFVRAQLAG KPFFWHIYPQ DENVHLDKLH301 AFWDKAHGFY TPETVSAHRR LSDDLNGGEA LSATQRLECW QTLQQHQNGW 351RQGAEDWSRY LFGQPSAPEK LAAFVSKHQK IR*w

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF32 shows 93.8% identity over a 81 as overlap with an ORF (ORF32a)from strain A of N. meningitidis:

The complete length ORF32a nucleotide sequence <SEQ ID 191> is:

1 ATGAATACTC CTCCTTTTTC TGCTGGANTT TTTTGCAAGG TCATCGACAA 51 TTTCGGCGACATCGGCGTTT CGTGGCGGCT TGCCCGTGTT TTGCACCGCG 101 AACTCGGTTG GCAGGTGCATTTGTGGACGG ACGATGTGTC CGCCTTGCGT 151 GCGCTTTGCC CTGATTTGCC CGATGTTCNCTGCGTTCATC AGGATATTCA 201 TGTCCGCACT TGGCATTCCG ATGCGGCAGA TATTGATACCGCGCCTGTTC 251 NCGATGTCGT CATCGAAACT TTTGCCTGCG ACCTGCCCGA AAATGTGCTG301 CACATCATCC GCCGACACAA GCCGCTTTGG CTGAANTGGG AATATTTGAG 351CGCGGAGGAN AGCAATGAAA GGCTGCACNT GATGCCTTCG CCGCAGGAGA 401 GTGTTCNAAAATANTTTTGG TTTATGGGTT TCAGCGAANN NAGCGGCGGA 451 CTGATACGCG AACGCGATTACTGCGAAGCC GTCCGTTTCG ATAGCGGAGC 501 CTTGCGCAAG AGGCTGATGC TTCCCGAAAAAAACGNCCCC GAATGGCTGC 551 TTTTCGGCTA TCGGAGCGAT GTTTGGGCAA AGTGGCTGGAAATGTGGCGA 601 CAGGCAGGCA GTCCGTTGAC ACTTTTGCTG GCNGGGGCGC ANATTATCGA651 CAGCCTCAAA CAAAACGGCG TTATTCCGCA AGATGCCCTG CAAAACGACG 701GCGATGTTTT TCAGACGGCA TCCGTCCGCC TCGTCAAAAT CCCTTTCGTG 751 CCGCAACAGGACTTCGACAA ACTGCTGCAC CTTGCCGACT GCGCCGTCAT 801 CCGCGGCGAA GACAGTTTCGTGCGCGCCCA GCTTGCGGGC AAACCCTTCT 851 TTTGGCACAT CTACCCGCAA GATGAGAATGTCCATCTCGA CAAACTCCAC 901 GCCTTTTGGG ATAAGGCACA CGGTTTCTAC ACGCCCGAAACCGCATCGGC 951 ACACCGCCGC CTTTCAGACG ACCTCAACGG CGGAGAGGCT TTATCCGCAA1001 CACAACGCCT CGAATGTTGG CAAATCCTGC AACAACATCA AAACGGCTGG 1051CGGCAAGGCG CGGAGGATTG GAGCCGTTAT CTTTTTGGGC AGCCTTCCGC 1101 ATCCGAAAAACTCGCCGCCT TTGTTTCAAA GCATCAAAAA ATACGCTAG

This encodes a protein having amino acid sequence <SEQ ID 192>:

1 MNTPPFSAGX FCKVIDNFGD IGVSWRLARV LHRELGWQVH LWTDDVSALR 51 ALCPDLPDVXCVHQDIHVRT WHSDAADIDT APVXDVVIET FACDLPENVL 101 HIIRRHKPLW LXWEYLSAEXSNERLHXMPS PQESVXKXFW FMGFSEXSGG 151 LIRERDYCEA VRFDSGALRK RLMLPEKNXPEWLLFGYRSD VWAKWLEMWR 201 QAGSPLTLLL AGAXIIDSLK QNGVIPQDAL QNDGDVFQTASVRLVKIPFV 251 PQQDFDKLLH LADCAVIRGE DSFVRAQLAG KPFFWHIYPQ DENVHLDKLH301 AFWDKAHGFY TPETASAHRR LSDDLNGGEA LSATQRLECW QILQQHQNGW 351RQGAEDWSRY LFGQPSASEK LAAFVSKHQK IR*

ORF32a and ORF32-1 show 93.2% identity in 382 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF32 shows 95.1% identity over a 82aa overlap with a predicted ORF(ORF32.ng) from N. gonorrhoeae:

An ORF32ng nucleotide sequence <SEQ ID 193> was predicted to encode aprotein having amino acid sequence <SEQ ID 194>:

1 MVMNTYAFPV CWIFCKVIDN FGDIGVSWRL ARVLHRELGW QVHLWTDDVS 51 ALRALCPDLPDVPFVHQDIH VRTWHSDAAD IDTAPVPDAV IETFACDLPE 101 NVLNIIRRHK PLWLNWEYLSAEESNERLHL MPSPQEGVQK YFWFMGFSEK 151 SGGLIRERDY REAVRFDTEA LRRRLVLPEKNAPEWLLFGY RGDVWAKWLD 201 MWQQAGSLMT LLLAGAQIID SLKQSGVIPQ NALQNEGGVFQTASVRLVKI 251 PFVPQQDFDK LLHLADCAVI RGEDSFVRTQ LAGKPFFWHI YPQDENVHLD301 KLHAFWDKAY GFYTPETASV HRLLSDDLNG GEALSATQRL ECGVL*

Further sequencing revealed the following DNA sequence <SEQ ID 195>:

1 ATGAATACAT ACGCTTTTCC TGTCTGTTGG ATTTTTTGCA AGGTCATCGA 51 CAATTTCGGCGACATCGGCG TTTCGTGGCG GCTCGCCCGT GTTTTGCACC 101 GCGAACTCGG TTGGCAGGTGCATTTGTGGA CGGACGACGT GTCCGCCTTG 151 CGCGCGCTTT GTCCCGATTT GCCCGATGTTCCCTTCGTTC ATCAGGATAT 201 TCATGTCCGC ACTTGGCATT CCGATGCGGC AGACATTGATACCGCGCCCG 251 TTCCCGATGC CGTTATCGAA ACTTTTGCCT GCGACCTGCC CGAAAATGTG301 CTGAACATCA TCCGCCGACA CAAACCGCTT TGGCTGAATT GGGAATATTT 351GAGCGCGGAG GAAAGCAATG AAAGGCTGCA CCTGATGCCT TCGCCGCAGG 401 AGGGCGTTCAAAAATATTTT TGGTTTATGG GTTTCAGCGA AAAAAGCGGC 451 GGGTTGATAC GCGAACGCGATTACCGCGAA GCCGTCCGTT TCGATACCGA 501 AGCCCTGCGC CGGCGGCTGG TGCTGCCCGAAAAAAACGCC CCCGAATGGC 551 TGCTTTTCGG CTATCGGGGC GATGTTTGGG CAAAGTGGCTGGACATGTGG 601 CAACAGGCAG GCAGCCTGAT GACCCTACTG CTGGCGGGGG CGCAAATTAT651 CGACAGCCTC AAACAAAGCG GCGTTATTCC GCAAAACGCC CTGCAAAAtg 701aaggcgGTGT CTTTCagacG gcatccgTcC gccttGTCAA AAtcCCGTTC 751 GTGCcGCAACAGGAcTTCGA CAAATTGCTG CAcctcgcCG ACTGCGCCGT 801 GATACGCGGC GAAGACAGTTTCGTGCGTAC CCAGCTTGCC GGAAAACCCT 851 TTTTTTGGCA CATCTACCCG CAAGACGAGAATGTCCATCT CGACAAACTC 901 CACGCCTTTT GGGATAAGGC ATACGGCTTC TACACGCCCGAAACCGCATC 951 GGTGCACCGC CTCCTTTCGG ACGACCTCAA CGGCGGAGAG GCTTTATCCG1001 CAACACAACG CCTCGAATGT TGGCAAACCC TGCAACAACA TCAAAACGGC 1051TGGCGGCAAG GCGCGGAGGA TTGGAGCCGT TATCTTTTCG GGCAGCCTTC 1101 CGCATCCGAAAAACTCGCCG CCTTTGTTTC AAAGCATCAA AAAATACGCT 1151 AG

This encodes a protein having amino acid sequence <SEQ ID 196;ORF32ng-1>:

1 MNTYAFPVCW IFCKVIDNFG DIGVSWRLAR VLHRELGWQV HLWTDDVSAL 51 RALCPDLPDVPFVHQDIHVR TWHSDAADID TAPVPDAVIE TFACDLPENV 101 LNIIRRHKPL WLNWEYLSAEESNERLHLMP SPQEGVQKYF WFMGFSEKSG 151 GLIRERDYRE AVRFDTEALR RRLVLPEKNAPEWLLFGYRG DVWAKWLDMW 201 QQAGSLMTLL LAGAQIIDSL KQSGVIPQNA LQNEGGVFQTASVRLVKIPF 251 VPQQDFDKLL HLADCAVIRG EDSFVRTQLA GKPFFWHIYP QDENVHLDKL301 HAFWDKAYGF YTPETASVHR LLSDDLNGGE ALSATQRLEC WQTLQQHQNG 351WRQGAEDWSR YLFGQPSASE KLAAFVSKHQ KIR*

ORF32ng-1 and ORF32-1 show 93.5% identity in 383 aa overlap:

On this basis, including the RGD sequence in the gonococcal protein,characteristic of adhesins, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF32-1 (42 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 7A shows the results ofaffinity purification of the His-fusion protein, and FIG. 7B shows theresults of expression of the GST-fusion in E. coli. Purified His-fusionprotein was used to immunise mice, whose sera were used for ELISA,giving a positive result. These experiments confirm that ORF32-1 is asurface-exposed protein, and that it is a useful immunogen.

Example 24

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 197>:

1 ..TTGTTCCTGC GTGTNAAAGT GGGGCGTTTT TTCAGCAGTC CGGCGACGTG 51  GTTTCGGGNC AAAGACCCTG TAAATCAGGC GGTGTTGCGG CTGTATNCGG 101  ACGAGTGGCG GCA.ACTTCG GTACGTTGGA AAATAGNCGC AACGTCGCAC 151  AGCCTGTGGC TCTGCACGCT GCTCGGAATG CTGGTGTCGG TATTGTTGCT 201  GCTTTTGGTG CGGCAATATA CGTTCAACTG GGAAAGCACG CTGTTGAGCA 251  ATGCCGCTTC GGTACGCGCG GTGGAAATGT TGGCATGGCT GCCGTCGAAA 301  CTCGGTTTCC CTGTCCCCGA TGCGCGGTCG GTCATCGAAG GCCGTCTGAA 351  CGGCAATATT GCCGATGCGC GGGCTTGGTC GGGGCTGCTG GTCGNCAGTA 401  TCGCCTGCTA NGGCATCCTG CCGCGCCTG..

This corresponds to the amino acid sequence <SEQ ID 198; ORF33>:

1 ..LFLRVKVGRF FSSPATWFRX KDPVNQAVLR LYXDEWRXTS VRWKIXATSH 51  SLWLCTLLGM LVSVLLLLLV RQYTFNWEST LLSNAASVRA VEMLAWLPSK 101  LGFPVPDARS VIEGRLNGNI ADARAWSGLL VXSIACXGIL PRL..

Further work revealed the complete nucleotide sequence <SEQ ID 199>:

1 ATGTTGAATC CATCCCGAAA ACTGGTTGAG CTGGTCCGTA TTTTGGACGA 51 AGGCGGTTTTATTTTCAGCG GCGATCCCGT ACAGGCGACG GAGGCTTTGC 101 GCCGCGTGGA CGGCAGTACGGAGGAAAAAA TCATCCGTCG GGCGGAGATG 151 ATTGACAGGA ACCGTATGCT GCGGGAGACGTTGGAACGTG TGCGTGCGGG 201 GTCGTTCTGG TTGTGGGTGG TGGCGGCGAC GTTTGCATTTTTTACCGGTT 251 TTTCAGTCAC TTATCTTCTA ATGGACAATC AGGGTCTGAA TTTCTTTTTG301 GTTTTGGCGG GCGTGTTGGG CATGAATACG CTGATGCTGG CAGTATGGTT 351GGCAATGTTG TTCCTGCGTG TGAAAGTGGG GCGTTTTTTC AGCAGTCCGG 401 CGACGTGGTTTCGGGGCAAA GACCCTGTAA ATCAGGCGGT GTTGCGGCTG 451 TATGCGGACG AGTGGCGGCAACCTTCGGTA CGTTGGAAAA TAGGCGCAAC 501 GTCGCACAGC CTGTGGCTCT GCACGCTGCTCGGAATGCTG GTGTCGGTAT 551 TGTTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGAAAGCACGCTG 601 TTGAGCAATG CCGCTTCGGT ACGCGCGGTG GAAATGTTGG CATGGCTGCC651 GTCGAAACTC GGTTTCCCTG TCCCCGATGC GCGGGCGGTC ATCGAAGGCC 701GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG GCTGCTGGTC 751 GGCAGTATCGCCTGCTACGG CATCCTGCCG CGCCTGCTGG CTTGGGTAGT 801 GTGTAAAATC CTTTTGAAAACAAGCGAAAA CGGATTGGAT TTGGAAAAGC 851 CCTATTATCA GGCGGTCATC CGCCGCTGGCAGAACAAAAT CACCGATGCG 901 GATACGCGTC GGGAAACCGT GTCCGCCGTT TCACCGAAAATCATCTTGAA 951 CGATGCGCCG AAATGGGCGG TCATGCTGGA GACCGAGTGG CAGGACGGCG1001 AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA GGGCGTTGCC 1051ACCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA AGCAGAAACC 1101 GGCGCAACTGCTTATCGGCG TGCGCGCCCA AACTGTGCCG GACCGCGGCG 1151 TGTTGCGGCA GATTGTCCGACTCTCGGAAG CGGCGCAGGG CGGCGCGGTG 1201 GTGCAGCTTT TGGCGGAACA GGGGCTTTCAGACGACCTTT CGGAAAAGCT 1251 GGAACATTGG CGTAACGCGC TGGCCGAATG CGGCGCGGCGTGGCTTGAGC 1301 CTGACAGGGC GGCGCAGGAA GGGCGTTTGA AAGACCAATA A

This corresponds to the amino acid sequence <SEQ ID 200; ORF33-1>:

1 MLNPSRKLVE LVRILDEGGF IFSGDPVQAT EALRRVDGST EEKIIRRAEM 51 IDRNRMLRETLERVRAGSFW LWVVAATFAF FTGFSVTYLL MDNQGLNFFL 101 VLAGVLGMNT LMLAVWLAMLFLRVKVGRFF SSPATWFRGK DPVNQAVLRL 151 YADEWRQPSV RWKIGATSHSLWLCTLLGML VSVLLLLLVR QYTFNWESTL 201 LSNAASVRAV EMLAWLPSKL GFPVPDARAVIEGRLNGNIA DARAWSGLLV 251 GSIACYGILP RLLAWVVCKI LLKTSENGLD LEKPYYQAVIRRWQNKITDA 301 DTRRETVSAV SPKIILNDAP KWAVMLETEW QDGEWFEGRL AQEWLDKGVA351 TNREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR LSEAAQGGAV 401VQLLAEQGLS DDLSEKLEHW RNALAECGAA WLEPDRAAQE GRLKDQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF33 shows 90.9% identity over a 143aa overlap with an ORF (ORF33a)from strain A of N. meningitidis:

The complete length ORF33a nucleotide sequence <SEQ ID 201> is:

1 ATGTTGAATC CATCCCGAAA ACTGGTTGAG CTGGTCCGTA TTTTGGAAGA 51 AGGCGGCTTTATTTTCAGCG GCGATCCCGT GCAGGCGACG GAGGCTTTGC 101 GCCGCGTGGA CGGCAGTACGGAGGAAAAAA TCATCCGTCG GGCGAAGATG 151 ATCGACAGGA ACCGTATGCT GCGGGAGACGTTGGAACGTG TGCGTGCGGG 201 GTCGTTCTGG TTGTGGGTGG CGGCGGCGAC GTTTGCGTTTNTTACCGNTT 251 TTTCAGTTAC TTATCTTCTA ATGGACAATC AGGGTCTGAA TTTCTTTTTG301 GTTTTGGCGG GCGTGNTGGG CATGAATACG CTGATGCTGG CAGTATGGTT 351GGCAATGTTG TTCCTGCGCG TGAAAGTGGG GCGTTTTTTC AGCAGTCCGG 401 CGACGTGGTTTCGGGGCAAA GACCCTGTCA ATCAGGCGGT GTTGCGGCTG 451 TATGCGGACG AGTGGCGGCNACCTTCGGTA CGTTGGAAAA TAGGCGCAAC 501 GTCGCACAGC CTGTGGCTCT GCACGCTGCTCGGAATGCTG GTGTCGGTAT 551 TGTTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGAAAGCACGCTG 601 TTGGGCGATT CGTCTTCGGT ACGGCTGGTG GAAATGTTGG CATGGCTGCC651 TGCGAAACTG GGTTTTCCCG TGCCTGATGC GCGGGCGGTC ATCGAAGGTC 701GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG GCTGCTGGTC 751 GGCAGTATCGCCTGCTACGG CATCCTGCCG CGCCTCTTGG CTTGGGCGGT 801 ATGCAAAATC CTTNTGNAAACAAGCGAAAA CGGCTTGGAT TTGGAAAAGC 851 NCNNNNNTCN NNCGNTCATC CGCCGCTGGCAGAACAAAAT CACCGATGCG 901 GATACGCGTC GGGAAACCGT GTCCGCCGTT TCGCCGAAAATCGTCTTGAA 951 CGATGCGCCG AAATGGGCGG TCATGCTGGA GACCGAATGG CAGGACGGCG1001 AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA GGGCGTTGCC 1051GCCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA AGCAGAAACC 1101 GGCGCAACTGCTTATCGGCG TGCGCGCCCA AACTGTGCCC GACCGCGGCG 1151 TGTTGCGGCA GATCGTCCGACTTTCGGAAG CGGCGCAGGG CGGCGCGGTG 1201 GTGCANCTTT TGGCGGAACA GGGGCTTTCAGACGACCTTT CGGAAAAGCT 1251 GGAACATTGG CGTAACGCGC TGACCGAATG CGGCGCGGCGTGGCTGGAAC 1301 CCGACAGAGC GGCGCAGGAA GGCCGTCTGA AAACCAACGA CCGCACTTGA

This encodes a protein having amino acid sequence <SEQ ID 202>:

1 MLNPSRKLVE LVRILEEGGF IFSGDPVQAT EALRRVDGST EEKIIRRAKM 51 IDRNRMLRETLERVRAGSFW LWVAAATFAF XTXFSVTYLL MDNQGLNFFL 101 VLAGVXGMNT LMLAVWLAMLFLRVKVGRFF SSPATWFRGK DPVNQAVLRL 151 YADEWRXPSV RWKIGATSHSLWLCTLLGML VSVLLLLLVR QYTFNWESTL 201 LGDSSSVRLV EMLAWLPAKL GFPVPDARAVIEGRLNGNIA DARAWSGLLV 251 GSIACYGILP RLLAWAVCKI LXXTSENGLD LEKXXXXXXIRRWQNKITDA 301 DTRRETVSAV SPKIVLNDAP KWAVMLETEW QDGEWFEGRL AQEWLDKGVA351 ANREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR LSEAAQGGAV 401VXLLAEQGLS DDLSEKLEHW RNALTECGAA WLEPDRAAQE GRLKTNDRT*

ORF33a and ORF33-1 show 94.1% identity in 444 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF33 shows 91.6% identity over a 143aa overlap with a predicted ORF(ORF33.ng) from N. gonorrhoeae:

An ORF33ng nucleotide sequence <SEQ ID 203> was predicted to encode aprotein having amino acid sequence <SEQ ID 204>:

1 MIDRDRMLRD TLERVRAGSF WLWVVVASMM FTAGFSGTYL LMDNQGLNFF 51LVLAGVLGMN TLMLAVWLAT LFLRVKVGRF FSSPATWFRG KGPVNQAVLR 101 LYADQWRQPSVRWKIGATAH SLWLCTLLGM LVSVLLLLLV RQYTFNWEST 151 LLSNAASVRA VEMLAWLPSKLGFPVPDARA VIEGRLNGNI ADARAWSGLL 201 VGSIVCYGIL PRLLAWVVCK ILLKTSENGLDLEKTYYQAV IRRWQNKITD 251 ADTRRETVSA VSPKIVLNDA PKWALMLETE WQDGQWFEGRLAQEWLDKGV 301 AANREQVAAL ETELKQKPAQ LLIGVRAQTV PDRGVLRQIV RLSEAAQGGA351 VVQLLAEQGL SDDLSEKLEH WRNALTECGA AWLEPDRVAQ EGRLKDQ*

Further sequence analysis revealed the following DNA sequence <SEQ ID205>:

1 ATGTTGaatC CATCCCgaAA ACTGgttgag ctGgTCCgtA Ttttgaataa 51 agggggtTTTattttcagcg gcgatcctgt gcaggcgacg gaggctttgc 101 gccgcgtgga cggcAGTACGGAggAaaaaa tcttccgtcg GGCGGAGAtg 151 atcgACAGGg accgtatgtt gcgggACaCgTtggaacGTG TGCGTGCggg 201 gtcgtTctgG TTATGGGTGG TggtggCAtC gATGATGTttaCCGCCGGAT 251 TTTCAGgcac ttatCttCTG ATGGACaatC AGGGGCtGAA TtTCTTTTTA301 GTTTTggcgG GAGTGTtggG CATGaatacG ctgATGCTGG CAGTATGGtt 351gGCAACGTTG TTCCTGCGCG TGAAAGTGGG ACGGTTTTTC AGCAGTCCGG 401 CGACGTGGTTTCGGGGCAAA GGCCCTGTAA ATCAGGCGGT GTTGCGGCTG 451 TATGCGGACC AGTGGCGGCAACCTTCGGTA CGATGGAAAA TAGGCGCAAC 501 GGCGCACAGC TTGTGGCTCT GCACGCTGCTCGGAATGCTG GTGTCGGTAT 551 TGCTGCTGCT TTTGGTGCGG CAATATACGT TCAACTGGGAAAGCACGCTG 601 TTGAGCAATG CCGCTTCGGT ACGCGCGGTG GAAATGTTGG CATGGCTGCC651 GTCGAAACTC GGTTTCCCTG TCCCCGATGC GCGGGCGGTC ATCGAAGGTC 701GTCTGAACGG CAATATTGCC GATGCGCGGG CTTGGTCGGG GCTGCTGGTC 751 GGCAGTATCGTCTGCTACGG CATCCTGCCG CGCCTCTTGG CTTGGGTAGT 801 GTGTAAAATC CTTTTGAAAACAAGCGAAAA CGGattgGAT TTGGAAAAAA 851 CCTATTATCA GGCGGTCATC CGCCGCTGGCAGAACAAAAT CACCGATGCG 901 GATACGCGTC GGGAAACCGT GTCCGCCGTT TCGCcgaAAATCGTCTTGAA 951 CGATGCGCCG AAATGGGCGC TCATGCTGGA GACCGAGTGG CAGGACGGCC1001 AATGGTTCGA GGGCAGGCTG GCGCAGGAAT GGCTGGATAA GGGCGTTGCC 1051GCCAATCGGG AACAGGTTGC CGCGCTGGAG ACAGAGCTGA AGCAGAAACC 1101 GGCGCAACTGCTTATCGGCG TACGCGCCCA AACTGTGCCG GACCGGGGCG 1151 TGCTGCGGCA GATTGTGCGGCTTTCGGAAG CGGCGCAGGG CGGCGCGGTG 1201 GTGCAGCTTT TGGCGGAACA GGGGCTTTCAGACGACCTTT CGGAAAAGCT 1251 GGAACATTGG CGTAACGCGC TGACCGAATG CGGCGCGGCGTGGCTTGAGC 1301 CTGACAGGGT GGCGCAGGAA GGCCGTTTGA AAGACCAATA A

This encodes a protein having amino acid sequence <SEQ ID 206;ORF33ng-1>:

1 MLNPSRKLVE LVRILNKGGF IFSGDPVQAT EALRRVDGST EEKIFRRAEM 51 IDRDRMLRDTLERVRAGSFW LWVVVASMMF TAGFSGTYLL MDNQGLNFFL 101 VLAGVLGMNT LMLAVWLATLFLRVKVGRFF SSPATWFRGK GPVNQAVLRL 151 YADQWRQPSV RWKIGATAHSLWLCTLLGML VSVLLLLLVR QYTFNWESTL 201 LSNAASVRAV EMLAWLPSKL GFPVPDARAVIEGRLNGNIA DARAWSGLLV 251 GSIVCYGILP RLLAWVVCKI LLKTSENGLD LEKTYYQAVIRRWQNKITDA 301 DTRRETVSAV SPKIVLNDAP KWALMLETEW QDGQWFEGRL AQEWLDKGVA351 ANREQVAALE TELKQKPAQL LIGVRAQTVP DRGVLRQIVR LSEAAQGGAV 401VQLLAEQGLS DDLSEKLEHW RNALTECGAA WLEPDRVAQE GRLKDQ*

ORF33ng-1 and ORF33-1 show 94.6% identity in 446 aa overlap:

Based on the presence of several putative transmembrane domains in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 25

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 207>:

1 ..CAGAAGAGTT TGTCGAGAAT TTCTTTATGG GGTTTGGGCG GCGTGTTTTT 51  CGGGGTGTCC GGTCTGGTAT GGTTTTCTTT GGGCGTTTCT TT.GAGTGCG 101  CCTGTTTTTC GGGTGTTTCT TTTCGGGGTT CGGGACGGGG GACGTTTGTG 151  GGCAGTACGG GGGTTTCTTT GAGTGTGTTT TCAGCTTGTG TTCC.GGCGT 201  CGTCCGGCTG CCTGTCGGTT TGAGCTGTGT CGGCAGGTTG CG..GTTTGA 251  CCCGGTTTTT CTTGGGTGCG GCAGGGGACG TCATTCTCCT GCCGCTTTCG 301  TCTGTGCCGT CCGGCTGTGC GGGTTCGGAT GAGGCGGCGT GGTGGTGTTC 351  GGGTTGGGCG GCATCTTGTT CCGACTACGC CGTTTGGCAG CCAGAATTCG 401  GTTTCGCGGG GGCTGTCGGT GTGTTGCGGT TCGGCTTGAA GGGTTTTGTC 451   GTCC..

This corresponds to the amino acid sequence <SEQ ID 208; ORF34>:

1 ..QKSLSRISLW GLGGVFFGVS GLVWFSLGVS XECACFSGVS FRGSGRGTFV 51  GSTGVSLSVF SACVXGVVRL PVGLSCVGRL XXLTRFFLGA AGDVILLPLS 101  SVPSGCAGSD EAAWWCSGWA ASCPTTPFGS QNSVSRGLSV CCGSA*RVLS 151   S..

Further work revealed the complete nucleotide sequence <SEQ ID 209>:

1 ATGATGATGC CGTTCATAAT GCTTCCTTGG ATTGCkGGTG TGCCTGCCGT 51 GCCGGGTCAGAATAGGTTGT CCAGAATTTC TTTATGGGGT TTGGGCGGCG 101 TGTTTTTCGG GGTGTCCGGTTTGGTATGGT TTTCTTTGGG CGTTTCTTTG 151 GGCTGCGCCT GTTTTTCGGG TGTTTCTTTTCGGGGTTCGG GACGGGGGAC 201 GTTTGTGGGC AGTACGGGGG TTTCTTTGAG TGTGTTTTCAGCTTGTGTTC 251 CGGCGTCGTC CGGCTGCCTG TCGGTTTGAG CTGTGTCGGC AGGTTGCGGT301 TTGACCCGGT TTTTCTTGGG TGCGGCAGGG GACGGCAGTC CGCTGCCGCT 351TTCGTCTGTG CCGTCCGGCT GTGCGGGTTC GGATGAGGCG GCGTGGTGGT 401 GTTCGGGTTGGGCGGCATCT TGTCCGACTA CGCCGTTTGG CAGCCAGAAT 451 TCGGTTTCGC GGGGGCTGTCGGTGTGTTGC GGTTCGGCTT GAAGGGTTTT 501 GTCGCCGTTC GGGTTGAATG TGCTGACGATGCCTATTGCC AATGCGCCGA 551 TGGCGGCGAT ACAGATGAGC AATACGGCGC GTATCAGGAGTTTGGGGGTC 601 AGCCTGAAGG GTTTGTTCGG TTTTTTTGCC ATTTTGATTG TGCTTTTGGG651 GTGTCGGGCA ATGCCGTCTG AAGGCGGTTC AGACGGCATT GCCGAGTCAG 701CGTTGGACGT AGTTTTGGTA GAGGGTGATG ACTTTTTGTA CGCCGACGGT 751 GGTGCTGACTTTTTGGGTAA TCTGCGCCTG TTCTTCGGGG GTGAGGATGC 801 CCATAACGTA GGTTACGTTGCCGTAGGTAA CGATTTTGAC GCGCGCCTGT 851 GTGGCGGGGC TGATGCCCAA CAGCGTGGCGCGGACTTTGG ATGTGTTCCA 901 AGTGTCGCCG GCGATGTCGC CGGCAGTGCG CGGCAGGGAGGCGACGGTAA 951 TATAGTTGTA CACGCCTTCG GCGGCCTGTT CGGAACGTGC AATCTGACCG1001 ACGAACTGTT TTTCGCCTTC GGTGGCGACT TGTCCGAGCA GCAGCAGGTG 1051GCGGTTGTAG CCGACGACGG AGATTTGGGG CGTGTAGCCT TTGGTTTGGT 1101 TGTTTTGGCGCAGATAGGAA CGGGCGGTGG TTTCGATACG CAACGCCATA 1151 ACGTTGTCGT CGGTTTGCGCGCCGGTGGTT CGGCGGTCGA CGGCGGATTT 1201 CGCGCCGACG GCGGCGCTTC CGATTACTGCGCTGACGCAG CCGCTAAGGG 1251 CAAGGCTGAA AATGGCGGCA ATCAGGGTGC GGACGGTGTGCGGTTTGGGT 1301 TTCATCGGGT GCTTCCTTTC TTGGGCGTTT CAGACGGCAT TGCTTTGCGC1351 CATGCCGTCT GA

This corresponds to the amino acid sequence <SEQ ID 210; ORF34-1>:

1 MMMPFIMLPW IAGVPAVPGQ NRLSRISLWG LGGVFFGVSG LVWFSLGVSL 51 GCACFSGVSFRGSGRGTFVG STGVSLSVFS ACVPASSGCL SV*AVSAGCG 101 LTRFFLGAAG DGSPLPLSSVPSGCAGSDEA AWWCSGWAAS CPTTPFGSQN 151 SVSRGLSVCC GSA*RVLSPF GLNVLTMPIANAPMAAIQMS NTARIRSLGV 201 SLKGLFGFFA ILIVLLGCRA MPSEGGSDGI AESALDVVLVEGDDFLYADG 251 GADFLGNLRL FFGGEDAHNV GYVAVGNDFD ARLCGGADAQ QRGADFGCVP301 SVAGDVAGSA RQGGDGNIVV HAFGGLFGTC NLTDELFFAF GGDLSEQQQV 351AVVADDGDLG RVAFGLVVLA QIGTGGGFDT QRHNVVVGLR AGGSAVDGGF 401 RADGGASDYCADAAAKGKAE NGGNQGADGV RFGFHRVLPF LGVSDGIALR 451 HAV*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF34 shows 73.3% identity over a 161aa overlap with an ORF (ORF34a)from strain A of N. meningitidis:

The complete length ORF34a nucleotide sequence <SEQ ID 211> is:

1 ATGATGATNC CGTTNATAAT GCTTCCTTGG ATTGCGGGTG TGCCTGCCGT 51 GCCGGGTCAGAAGAGGTTGT CGAGAANTTC TTTATGGGGT TTAGGCGGCN 101 TGTTTTTCGG GGTGTCCGGTTTGGTATGGT TTTCTTTGGG CGTTTCTNTT 151 TCTTTGGGTG TTTCTNTGGG CTGTGCCTGTTTTTCGGGTG TTTCTTTTCG 201 GGGTTCGGGA CGGGGGACGT TTGTGGGCAG TACNGGGGTTTCTTTGAGTG 251 TGTTTTCAGC TTGTGCTCCG GCGTCGTCCG GCTGCCTGTC GGTTTNAGCT301 GTGTCGGCAG GTTGCGGTTT GACCCGGNTT TTCTTNGGTG CGGCAGGGGA 351CGGCAGTCCG CTGCCGCTTT CGTCTGTGCC GTCCGGCTGT GCGGGTGCGG 401 ATGAGGAGGCGTNGTNGTGT TCGGGTTGGG CGGCATCTTG TCCGACTACG 451 CCGTTTGGCA GCCAGAATTCGGTTTCGCGG GGGCTGTCGG TGTGTTGCGG 501 TTCGGTNTGG AGGGTTTTGT CNCCGTTCGGGTNGAATGTG CTGACGATGC 551 CTATTGCCAA TGCGCCGATG GCGGTGATAC AGATGAGCAATACGGCGCGT 601 ATCAGGAGTT TGGGGGTCAG CCTGAAGGGT TTGTTCNGTT TTTTTGCCAT651 TTTGATTGTG CTTTTGGGGT GTCGGGCAAT GCCGTCTGAA GGCGGTTCAG 701ACGGCATTGC CGAGTCAGCG TTGGACGTAG TTTNGGTAGA GGGTGATGAC 751 TTTTTGTACGCCGACGGTGG TGCTGACTTT TTGGGTAATC TGCGCCTGTT 801 CTTCGGGGGT GAGGATGCCCATAACGTAGG TTACGTTGCC GTAGGTAACG 851 ATTTTGACGC GCGCCTGTGT GGCGGGGCTGATGCCCAACA GCGTGGCGCG 901 GACTTTGGAT GTGTTCCAAG TGTCGCCGGC GATGTCGCCGGCAGTGCGCG 951 GCAGGGAGGC GACGGTAATG TANTTGTACA CGCCTTCGGC GGCCTGTTCG1001 GAACGTGCAA TCTGACCGAC GAACTGTTTC TCGCCTTCGG TGGCGACTTG 1051TCCGAGCAGC AGCAGGTGGC GGTTGTAGCC GACAACGGAG ATTTGGGGCG 1101 TGTANCCTTTGGTTTGGTTG TTTTGGCGCA GATAGGAGCG GGCGGTGGTT 1151 TCGATACGCA GCGCCATTACGTTGTCGTCG GTTNGCGCGC CGGTGGTTCG 1201 GCGGTCGACG GCGGATTTCG CGCCGACCGCCGCGCCGCCG ACGACTGCGC 1251 TGACGCAGCC GCCGAGGGCA AGGCTGAGGA CGGCGGCAGTCAGGGTGCGG 1301 ACGGTGTGCG GTTTGGGTTT CATCGGGTGC TTCCTTTCTT GGGCGTTTCA1351 GACGGCATTG CTTTGCGCCA TGCCGTCTGA

This encodes a protein having amino acid sequence <SEQ ID 212>:

1 MMXPXIMLPW IAGVPAVPGQ KRLSRXSLWG LGGXFFGVSG LVWFSLGVSX 51SLGVSXGCAC FSGVSFRGSG RGTFVGSTGV SLSVFSACAP ASSGCLSVXA 101 VSAGCGLTRXFXGAAGDGSP LPLSSVPSGC AGADEEAXXC SGWAASCPTT 151 PFGSQNSVSR GLSVCCGSVWRVLSPFGXNV LTMPIANAPM AVIQMSNTAR 201 IRSLGVSLKG LFXFFAILIV LLGCRAMPSEGGSDGIAESA LDVVXVEGDD 251 FLYADGGADF LGNLRLFFGG EDAHNVGYVA VGNDFDARLCGGADAQQRGA 301 DFGCVPSVAG DVAGSARQGG DGNVXVHAFG GLFGTCNLTD ELFLAFGGDL351 SEQQQVAVVA DNGDLGRVXF GLVVLAQIGA GGGFDTQRHY VVVGXRAGGS 401AVDGGFRADR RAADDCADAA AEGKAEDGGS QGADGVRFGF HRVLPFLGVS 451 DGIALRHAV*

ORF34a and ORF34-1 show 91.3% identity in 459 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF34 shows 77.6% identity over a 161 as overlap with a predicted ORF(ORF34.ng) from N. gonorrhoeae:

The complete length ORF34ng nucleotide sequence <SEQ ID 213> is:

1 ATGATGATGC CGTTCATAAT GCTTCCTTGG ATTGCGGGTG TGCCTGCCGT 51 GCCGGGTCAAAAGAGGTTGT CGAGAATCTC TTTATGGGGT TTGGCCGGCG 101 TGTTTTTCGG GGTGTCCGGTTTGGTATGGT TTTCTTTGGG CGTTTCTTTT 151 TCTTTGGGTG TTTCTTTGGG CTGCGCCTGTTTTTCGGGTG TTTCTTTTCG 201 GGGTTCGGGA TGGGGGGCGT TTGTGGGCAG TACGGGGGTTTCTTTGAGTG 251 TGTTTTCAGC TTGTGTTCCG GTGCCGGTTA ACGAATCGGC TGCCCGGGCC301 GCATCCGAAG GGCGCGGTTT gACCCGGTTT TTCTTGGGTG CGGCAGGGGA 351CGGCAGTCCG CTGCCGCTTT CTTCTGTGCC GTCCGGCTGT GCGGGTTCGG 401 ATGAGGCGGCGTGGTGGTGT TCGGGTTGGG CGGCATCTTG TCCGACGGCG 451 CCGTTTGGCA GCCAGAATTCGGTTTCGCGG GGGCTGTCGG TGTGTTGCGG 501 TTCGGTTTGG AGGGTTTTGT CGCCGTTCGGGTTGAATGTG CTGACGATGC 551 CTACTGCCAA TGCGCCGATG GCGGTGATAC AGATGAGCAATACGGCGCGT 601 ATCAGGAGTT TGGGGGTCAG CCTGAAGGGT TTGTTCGGTT TTTTTGCCAT651 TTTGATTGTG CTTTTGGGGT GTCGGGCAAT GCCGTCTGAA GGCGGTTCAG 701ACGGCATTGC CGAGTCAGCG TTGGACGTAG TTTTGGTAGA GGGTAATGAC 751 TTTTTGTACGCCGAcggTGG TGCTGACTTT TTGGGTAATC TGCGCCTGTT 801 CTTCGGGGGT GAGGATGCCCATAACGTAGG TTACATTGCC GTAGGTAATG 851 ATTTTGACGC GCGCCTGTGT AGCGGGGCTGATGCCCAGCA GcgtgGCGCG 901 GACTTTGGAC GTGTTCCAAG TGTCGCCGGC GATGTCGCCCGCAGTGCGCG 951 GCAGGGAGGC GACGGTAATG TAGTTGTATA CGCCTTCGGC GGCCTGTTCG1001 GAACGTGCAA TCTGACCGAC GAACTGTTTT TCGCCTTCGG TGGCGACTTG 1051TCCGAGCAGC AGCAGGTGGC GGTTGTAGCC GACGACGGAG ATTTGGGGCG 1101 TGTAGCCTTTGGTTTGGTTG TTTTGGCGCA GGTAGGAACG GGCGGTGGTT 1151 TCGATACGCA ACGCCATAACGTtgtCATCG GTTtgcgcgc CGGTGGTTcg 1201 gCGGTCGATG ACGGATTTTG CGCCGACGGCGGCCCCGCCG ACGACTGCGC 1251 TGAAGCAGCC GCCGAGGGCA AGGCTGAGGA CGGCGGCAATCAGGGTGCGG 1301 ACGGTGTGTG GTTTGGGTTT CATCGGGGAC TTCCTTTCTT GGGCGTTTCA1351 GACGGCATTG CTTTGCGCCA TGCCGTCTGA

This encodes a protein having amino acid sequence <SEQ ID 214>:

1 MMMPFIMLPW IAGVPAVPGQ KRLSRISLWG LAGVFFGVSG LVWFSLGVSF 51SLGVSLGCAC FSGVSFRGSG WGAFVGSTGV SLSVFSACVP VPVNESAARA 101 ASEGRGLTRFFLGAAGDGSP LPLSSVPSGC AGSDEAAWWC SGWAASCPTA 151 PFGSQNSVSR GLSVCCGSVWRVLSPFGLNV LTMPTANAPM AVIQMSNTAR 201 IRSLGVSLKG LFGFFAILIV LLGCRAMPSEGGSDGIAESA LDVVLVEGND 251 FLYADGGADF LGNLRLFFGG EDAHNVGYIA VGNDFDARLCSGADAQQRGA 301 DFGRVPSVAG DVARSARQGG DGNVVVYAFG GLFGTCNLTD ELFFAFGGDL351 SEQQQVAVVA DDGDLGRVAF GLVVLAQVGT GGGFDTQRHN VVIGLRAGGS 401AVDDGFCADG GPADDCAEAA AEGKAEDGGN QGADGVWFGF HRGLPFLGVS 451 DGIALRHAV*

ORF34ng and ORF34-1 show 90.0% identity in 459 aa overlap:

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 26

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 215>:

1 ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG CGCTCATCCT 51 CGCCGCCTGCGGATT.CAAA AAGACAGCGC GCCCGCCGCA TCCGCTTCTG 101 CCGCCGCCGA CAACGGCGCGGCGTAAAAAA GAAATCGTCT TCGGCACGAC 151 CGTCGGCGAC TTCGGCGATA TGGTCAAAGAACAAATCCAA GCCGAGCTGG 201 AGAAAAAAGG CTACACCGTC AAACTGGTCG AGTTTACCGACTATGTACGC 251 CCGAATCTGG CATTGGCTGA GGGCGAGTTG

This corresponds to the amino acid sequence <SEQ ID 216; ORF4>:

1 MKTFFKTLSA AALALILAAC G.QKDSAPAA SASAAADNGA AKKEIVFGTT 51 VGDFGDMVKEQIQAELEKKG YTVKLVEFTD YVRPNLALAE GEL

Further sequence analysis revealed the complete nucleotide sequence <SEQID 217>:

1 ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG CGCTCATCCT 51 CGCCGCCTGCGGCGGTCAAA AAGACAGCGC GCCCGCCGCA TCCGCTTCTG 101 CCGCCGCCGA CAACGGCGCGGCGAAAAAAG AAATCGTCTT CGGCACGACC 151 GTCGGCGACT TCGGCGATAT GGTCAAAGAACAAATCCAAG CCGAGCTGGA 201 GAAAAAAGGC TACACCGTCA AACTGGTCGA GTTTACCGACTATGTACGCC 251 CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT CTTCCAACAC301 AAACCCTATC TTGACGACTT CAAAAAAGAA CACAATCTGG ACATCACCGA 351AGTCTTCCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG GGCAAGCTGA 401 AATCGCTGGAAGAAGTCAAA GACGGCAGCA CCGTATCCGC GCCCAACGAC 451 CCGTCCAACT TCGCCCGCGTCTTGGTGATG CTCGACGAAC TGGGTTGGAT 501 CAAACTCAAA GACGGCATCA ATCCGTTGACCGCATCCAAA GCGGACATCG 551 CCGAGAACCT GAAAAACATC AAAATCGTCG AGCTTGAAGCCGCGCAACTG 601 CCGCGTAGCC GCGCCGACGT GGATTTTGCC GTCGTCAACG GCAACTACGC651 CATAAGCAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA GAACCGAGCT 701TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA AGACAGCCAA 751 TGGCTTAAAGACGTAACCGA GGCCTATAAC TCCGACGCGT TCAAAGCCTA 801 CGCGCACAAA CGCTTCGAGGGCTACAAATC CCCTGCCGCA TGGAATGAAG 851 GCGCAGCCAA ATAA

This corresponds to the amino acid sequence <SEQ ID 218; ORF4-1>:

1 MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA AKKEIVFGTT 51 VGDFGDMVKEQIQAELEKKG YTVKLVEFTD YVRPNLALAE GELDINVFQH 101 KPYLDDFKKE HNLDITEVFQVPTAPLGLYP GKLKSLEEVK DGSTVSAPND 151 PSNFARVLVM LDELGWIKLK DGINPLTASKADIAENLKNI KIVELEAAQL 201 PRSRADVDFA VVNGNYAISS GMKLTEALFQ EPSFAYVNWSAVKTADKDSQ 251 WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF4 shows 93.5% identity over a 93aa overlap with an ORF (ORF4a) fromstrain A of N. meningitidis:

The complete length ORF4a nucleotide sequence <SEQ ID 219> is:

1 ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG CGCTCATCCT 51 CGCCGCCTGCGGCGGTCAAA AAGATAGCGC GCCCGCCGCA TCCGCTTCTG 101 CCGCCGCCGA CAACGGCGCGGCGAANAAAG AAATCGTCTT CGGCACGACC 151 GTCGGCGACT TCGGCGATAT GGTCAAAGAACANATCCAAC CCGAGCTGGA 201 GAAAAAAGGC TACACCGTCA AACTGGTCGA GTNTACCGACTATGTGCGCN 251 CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT CTTNCAACAC301 ANACNCTATC TTGACGACTN CAAAAAANAA CACAATCTGG ACATCACCNN 351AGTCTTNCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG GGCAAGCTGA 401 AATCGCTGGANNAAGTCAAA GANGGCAGCA CCGTATCCGC GCCCAACGAC 451 CCGTNNNACT TCGNCCGCGTCTTGGTGATG CTCGACGAAC TGGGTTNGAT 501 CAAACTCAAA GACNGCATCA NNNNGNNGNNNNNANCNANA NNNGANANNN 551 NNNNANNNNT NNNNNNNNNN NNNNNCNNCG NNNNNNNANNNNNNNNNNNN 601 NCGNNTNNNN NNGCNNNNNT NNANNNTNNN NNCNNCNNNN NNNNNTNNNN651 NANNANNAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA GAACCGAGCT 701TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA AGACAGCCAA 751 TGGCTTAAAGACGTAACCGA GGCCTATAAC TCCGACGCGT TCAAAGCCTA 801 CGCGCACAAA CGCTTCGAGGGCTACAAATC CCCTGCCGCA TGGAATGAAG 851 GCGCAGCCAA ATAA

This is predicted to encode a protein having amino acid sequence <SEQ ID220>:

1 MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA AXKEIVFGTT 51 VGDFGDMVKEXIQPELEKKG YTVKLVEXTD YVRXNLALAE GELDINVXQH 101 XXYLDDXKKX HNLDITXVXQVPTAPLGLYP GKLKSLXXVK XGSTVSAPND 151 PXXFXRVLVM LDELGXIKLK DXIXXXXXXXXXXXXXXXXX XXXXXXXXXX 201 XXXXAXXXXX XXXXXXXXXS GMKLTEALFQ EPSFAYVNWSAVKTADKDSQ 251 WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

A leader peptide is underlined.

Further analysis of these strain A sequences revealed the complete DNAsequence <SEQ ID 221>:

1 ATGAAAACCT TCTTCAAAAC CCTTTCCGCC GCCGCACTCG CGCTCATCCT 51 CGCCGCCTGCGGCGGTCAAA AAGATAGCGC GCCCGCCGCA TCCGCTTCTG 101 CCGCCGCCGA CAACGGCGCGGCGAAAAAAG AAATCGTCTT CGGCACGACC 151 GTCGGCGACT TCGGCGATAT GGTCAAAGAACAAATCCAAC CCGAGCTGGA 201 GAAAAAAGGC TACACCGTCA AACTGGTCGA GTTTACCGACTATGTGCGCC 251 CGAATCTGGC ATTGGCTGAG GGCGAGTTGG ACATCAACGT CTTCCAACAC301 AAACCCTATC TTGACGACTT CAAAAAAGAA CACAATCTGG ACATCACCGA 351AGTCTTCCAA GTGCCGACCG CGCCTTTGGG ACTGTACCCG GGCAAGCTGA 401 AATCGCTGGAAGAAGTCAAA GACGGCAGCA CCGTATCCGC GCCCAACGAC 451 CCGTCCAACT TCGCCCGCGTCTTGGTGATG CTCGACGAAC TGGGTTGGAT 501 CAAACTCAAA GACGGCATCA ATCCGCTGACCGCATCCAAA GCGGACATTG 551 CCGAAAACCT GAAAAACATC AAAATCGTCG AGCTTGAAGCCGCGCAACTG 601 CCGCGTAGCC GCGCCGACGT GGATTTTGCC GTCGTCAACG GCAACTACGC651 CATAAGCAGC GGCATGAAGC TGACCGAAGC CCTGTTCCAA GAACCGAGCT 701TTGCCTATGT CAACTGGTCT GCCGTCAAAA CCGCCGACAA AGACAGCCAA 751 TGGCTTAAAGACGTAACCGA GGCCTATAAC TCCGACGCGT TCAAAGCCTA 801 CGCGCACAAA CGCTTCGAGGGCTACAAATC CCCTGCCGCA TGGAATGAAG 851 GCGCAGCCAA ATAA

This encodes a protein having amino acid sequence <SEQ ID 222; ORF4a-1>:

1 MKTFFKTLSA AALALILAAC GGQKDSAPAA SASAAADNGA AKKEIVFGTT 51 VGDFGDMVKEQIQPELEKKG YTVKLVEFTD YVRPNLALAE GELDINVFQH 101 KPYLDDFKKE HNLDITEVFQVPTAPLGLYP GKLKSLEEVK DGSTVSAPND 151 PSNFARVLVM LDELGWIKLK DGINPLTASKADIAENLKNI KIVELEAAQL 201 PRSRADVDFA VVNGNYAISS GMKLTEALFQ EPSFAYVNWSAVKTADKDSQ 251 WLKDVTEAYN SDAFKAYAHK RFEGYKSPAA WNEGAAK*

ORF4a-1 and ORF4-1 show 99.7% identity in 287 aa overlap:

Homology with an Outer Membrane Protein of Pasteurella haemolitica(Accession q08869).

ORF4 and this outer membrane protein show 33% aa identity in 91aaoverlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF4 shows 93.6% identity over a 94aa overlap with a predicted ORF(ORF4.ng) from N. gonorrhoeae:

The complete length ORF4ng nucleotide sequence <SEQ ID 223> waspredicted to encode a protein having amino acid sequence <SEQ ID 224>:

1 MKTFFKTLST ASLALILAAC GGQKDSAPAA SAAAPSADNG AAKKEIVFGT 51 TVGDFGDMVKEQIQAELEKK GYTVKLVEFT DYVRPNLALA EGELDINVFQ 101 HKPYLDDFKK EHNLDITEAFQVPTAPLGLY PGKLKSLEEV KDGSTVSAPN 151 DPSNFARALV MLNELGWIKL KDGINPLTASKADIAENLKN IKIVELEAAQ 201 LPRSRADVDF AVVNGNYAIS SGMKLTEALF QEPSFAYVNWSAVKTADKDS 251 QWLKDVTEAY NSDAFKAYAH KRFEGYKYPA AWNEGAAK*

Further analysis revealed the complete length ORF4ng DNA sequence <SEQID 225> to be:

1 atgAAAACCT TCTTCAAAAC cctttccgcc gccgcaCTCG CGCTCATCCT 51 CGCAGCCTGcggCggtcaAA AAGACAGCGC GCCCgcagcc tctgcCGCCG 101 CCCCTTCTGC CGATAACGgcgCgGCGAAAA AAGAAAtcgt ctTCGGCACG 151 Accgtgggcg acttcggcgA TAtggTCAAAGAACAAATCC AagcCGAgct 201 gGAGAAAAAA GgctACACcg tcAAattggt cgaatttaccgactatgtGC 251 gCCCGAATCT GGCATTGGCG GAGGGCGAGT TGGACATCAA CGTCTTCCAA301 CACAAACCCT ATCTTGACGA TTTCAAAAAA GAACACAACC TGGACATCAC 351CGAAGCCTTC CAAGTGCCGA CCGCGCCTTT GGGACTGTAT CCGGGCAAAC 401 TGAAATCGCTGGAAGAAGTC AAAGACGGCA GCACCGTATC CGCGCCCAac 451 gACccgTCCA ACTTCGCACGCGCCTTGGTG ATGCTGAACG AACTGGGTTG 501 GATCAAACTC AAAGACGGCA TCAATCCGCTGACCGCATCC AAAGCCGACA 551 TCGCGGAAAA CCTGAAAAAC ATCAAAATCG TCGAGCTTGAAGCCGCACAA 601 CTGCCGCGCA GCCGCGCCGA CGTGGATTTT GCCGTCGTCA ACGGCAACTA651 CGCCATAAGC AGCGGCATGA AGCTGACCGA AGCCCTGTTC CAAGAGCCGA 701GCTTTGCCTA TGTCAACTGG TCTGCCgtcA AAACCGCCGA CAAAGACAGC 751 CAATGGCTTAAAGACGTAAC CGAGGCCTAT AACTCCGACG CGTTCAAAGC 801 CTACGCGCAC AAACGCTTCGAGGGCTACAA ATACCCTGCC GCATGGAATG 851 AAGGCGCAGC CAAATAA

This encodes a protein having amino acid sequence <SEQ ID 226;ORF4ng-1>:

1 MKTFFKTLSA AALALILAAC GGQKDSAPAA SAAAPSADNG AAKKEIVFGT 51 TVGDFGDMVKEQIQAELEKK GYTVKLVEFT DYVRPNLALA EGELDINVFQ 101 HKPYLDDFKK EHNLDITEAFQVPTAPLGLY PGKLKSLEEV KDGSTVSAPN 151 DPSNFARALV MLNELGWIKL KDGINPLTASKADIAENLKN IKIVELEAAQ 201 LPRSRADVDF AVVNGNYAIS SGMKLTEALF QEPSFAYVNWSAVKTADKDS 251 QWLKDVTEAY NSDAFKAYAH KRFEGYKYPA AWNEGAAK*

This shows 97.6% identity in 288 aa overlap with ORF4-1:

In addition, ORF4ng-1 shows significant homology with an outer membraneprotein from the database:

Based on this analysis, including the homology with the outer membraneprotein of Pasteurella haemolitica, and on the presence of a putativeprokaryotic membrane lipoprotein lipid attachment site in the gonococcalprotein, it was predicted that these proteins from N. meningitidis andN. gonorrhoeae, and their epitopes, could be useful antigens forvaccines or diagnostics, or for raising antibodies.

ORF4-1 (30 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIGS. 8A and 8B show,respectively, the results of affinity purification of the His-fusion andGST-fusion proteins. Purified His-fusion protein was used to immunisemice, whose sera were used for ELISA (positive result), Western blot(FIG. 8C), FACS analysis (FIG. 8D), and a bactericidal assay (FIG. 8E).These experiments confirm that ORF4-1 is a surface-exposed protein, andthat it is a useful immunogen.

FIG. 8F shows plots of hydrophilicity, antigenic index, and AMPHIregions for ORF4-1.

Example 27

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 227>:

1 CCTCGTCGTC CTCGGCATGC TCCAGTTTCA AGGGGCGATT TACTCCAAGG 51 CGGTGGAACGTATGCTCGGC ACGGTCATCG GGCTGGGCGC GGGTTTGGGC 101 GTTTTATGGC TGAACCAGCATTATTTCCAC GGCAACCTCC TCTTCTACCT 151 CACCGTCGGC ACGGCAAGCG CACTGGCCGGCTGGGCGGCG GTCGGCAAAA 201 ACGGCTACGT CCCTmTGCTG GCAGGGCTGA CGATGTGTATGCTCATCGGC 251 GACAACGGCA GCGAATGGCT CGACAGCGGA CTCATGCGCG CCATGAACGT301 CCTCATCGGC GyGGCCATCG CCATCGCCGC CGCCAAACTG CTGCCGCTGA 351AATCCACACT GATGTGGCGT TTCATGCTTG CCGACAACCT GGCCGACTGC 401 AGCAAAATGATTGCCGAAAT CAGCAACGGC AGGCGCATGA CCCGCGAACG 451 CCTCGAGGAG AACATGGCGAAAATGCGCCA AATCAACGCA CGCATGGTCA 501 AAAGCCGCAG CCATCTCGCC GCCACATCGGGCGAAAGCTG CATCAGCCCC 551 GCCATGATGG AAGCCATGCA GCACGCCCAC CGTAAAATCGTCAACACCAC 601 CGAGCTGCTC CTGACCACCG CCGCCAAGCT GCAATCTCCC AAACTCAACG651 GCAGCGAAAT CCGGCTGCTT GACCGCCACT TCACACTGCT CCAAAC.... 701............................. GC AGACACGCCC GCCGCATCCG 751 CATCGACACCGCCATCAACC CCGAACTGGA AGCCCTCGCC GAACACCTCC 801 ACTACCAATG GCAGGGCTTCCTCTGGCTCA GCACCGATAT GCGTCAGGAA 851 ATTTCCGCCC TCGTCATCCT GCTGCAACGCACCCGCCGCA AATGGCTGGA 901 TGCCCACGAA CGCCAACACC TGCGCCAAAG CCTGCTTGA

This corresponds to the amino acid sequence <SEQ ID 228; ORF8>:

1 ......PRRP RHAPVSRGDL LQGGGTYARH GHRAGRGFGR FMAEPALFPR 51 QPPLLPHRRHGKRTGRLGGG RQKRLRPXAG RADDVYAHRR QRQRMARQRT 101 HARHERPHRR GHRHRRRQTAAAEIHTDVAF HACRQPGRLQ QNDCRNQQRQ 151 AHDPRTPRGE HGENAPNQRT HGQKPQPSRRHIGRKLHQPR HDGSHAARPP 201 XNRQHHRAAP DHRRQAAISQ TQRQRNPAAX PPLHTAPN...........Q 251 TRPPHPHRHR HQPRTGSPRR TPPLPMAGLP LAQHRYASGN FRPRHPAATH301 PPQMAGCPRT PTPAPKPA*

Computer analysis of this amino acid sequence gave the followingresults:

Sequence Motifs

ORF8 is proline-rich and has a distribution of proline residuesconsistent with a surface localization. Furthermore the presence of anRGD motif may indicate a possible role in bacterial adhesion events.

Homology with a Predicted ORF from N. gonorrhoeae

ORF8 shows 86.5% identity over a 312aa overlap with a predicted ORF(ORF8.ng) from N. gonorrhoeae:

The complete length ORF8ng nucleotide sequence <SEQ ID 229> is predictedto encode a protein having amino acid sequence <SEQ ID 230>:

1 MDRDDRLRRP RHAPVPRRDL LQRGGTYARY GHRAGRGFGR FMAEPALFPR 51 QPPLLPDHRHGKRTGRLGGG RQKRLRPYVG GADDVHAHRR QRQRMARQRP 101 DARDERPHRR RHRHCRRQTAAAEIHTDVAF HACRQPGRLQ QNDCRNQQRQ 151 AYDARTFGAE YGQNAPNQRT HGQKPQPPRRHIGRKPHQPL HDGSHAARPP 201 QNRQHHRAAP DHRRQAAISQ TQRQRNPAAR PPLHTAPNRPATNRRPHQRQ 251 TRPPHPHRHR HQPRTGSPRR TPPLPMAGFP LAQHQYASGN FRPRHPPATH301 PPQMAGCPRT PTPAPKPA*

Based on the sequence motifs in these proteins, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 28

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 231>:

1 ..GAAATCAGCC TGCGGTCCGA CNACAGGCCG GTTTCCGTGN CGAAGCGGCG 51  GGATTCGGAA CGTTTTCTGC TGTTGGACGG CGGCAACAGC CGGCTCAAGT 101  GGGCGTGGGT GGAAAACGGC ACGTTCGCAA CCGTCGGTAG CGCGCCGTAC 151  CGCGATTTGT CGCCTTTGGG CGCGGAGTGG GCGGAAAAGG CGGATGGAAA 201  TGTCCGCATC GTCGGTTGCG CTGTGTGCGG AGAATTCAAA AAGGCACAAG 251  TGCAGGAACA GCTCGCCCGA AAAATCGAGT GGCTGCCGTC TTCCGCACAG 301  GCTTT.GGCA TACGCAACCA CTACCGCCAC CCCGAAGAAC ACGGTTCCGA 351  CCGCTGGTTC AACGCCTTGG GCAGCCGCCG CTTCAGCCGC AACGCCTGCG 401  TCGTCGTCAG TTGCGGCACG GCGGTAACGG TTGACGCGCT CACCGATGAC 451  GGACATTATC TCGGAGA.GG AACCATCATG CCCGGTTTCC ACCTGATGAA 501  AGAATCGCTC GCCGTCCGAA CCGCCAACCT CAACCGGCAC GCCGGTAAGC 551  GTTATCCTTT CCCGACCGG..

This corresponds to the amino acid sequence <SEQ ID 232; ORF61>:

1 ..EISLRSDXRP VSVXKRRDSE RFLLLDGGNS RLKWAWVENG TFATVGSAPY 51  RDLSPLGAEW AEKADGNVRI VGCAVCGEFK KAQVQEQLAR KIEWLPSSAQ 101  AXGIRNHYRH PEEHGSDRWF NALGSRRFSR NACVVVSCGT AVTVDALTDD 151  GHYLGXGTIM PGFHLMKESL AVRTANLNRH AGKRYPFPT..

Further work revealed the complete nucleotide sequence <SEQ ID 233>:

1 ATGACGGTTT TGAAGCTTTC GCACTGGCGG GTGTTGGCGG AGCTTGCCGA 51 CGGTTTGCCGCAACACGTCT CGCAACTGGC GCGTATGGCG GATATGAAGC 101 CGCAGCAGCT CAACGGTTTTTGGCAGCAGA TGCCGGCGCA CATACGCGGG 151 CTGTTGCGCC AACACGACGG CTATTGGCGGCTGGTGCGCC CATTGGCGGT 201 TTTCGATGCC GAAGGTTTGC GCGAGCTGGG GGAAAGGTCGGGTTTTCAGA 251 CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT ACTGGAATTG301 GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGCG TGACCCACCT 351GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG CACCGTTTGG 401 GCGAGTGTCTGATGTTCAGT TTTGGCTGGG TGTTTGACCG GCCGCAGTAT 451 GAGTTGGGTT CGCTGTCGCCTGTTGCGGCA GTGGCGTGTC GGCGCGCCTT 501 GTCGCGTTTA GGTTTGGATG TGCAGATTAAGTGGCCCAAT GATTTGGTTG 551 TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACGGTCAGGACGGGC 601 GGCAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTTG TCCTGCCCAA651 GGAAGTAGAA AATGCCGCTT CCGTGCAATC GCTGTTTCAG ACGGCATCGC 701GGCGGGGCAA TGCCGATGCC GCCGTGCTGC TGGAAACGCT GTTGGTGGAA 751 CTGGACGCGGTGTTGTTGCA ATATGCGCGG GACGGATTTG CGCCTTTTGT 801 GGCGGAATAT CAGGCTGCCAACCGCGACCA CGGCAAGGCG GTATTGCTGT 851 TGCGCGACGG CGAAACCGTG TTCGAAGGCACGGTTAAAGG CGTGGACGGA 901 CAAGGCGTTT TGCACTTGGA AACGGCAGAG GGCAAACAGACGGTCGTCAG 951 CGGCGAAATC AGCCTGCGGT CCGACGACAG GCCGGTTTCC GTGCCGAAGC1001 GGCGGGATTC GGAACGTTTT CTGCTGTTGG ACGGCGGCAA CAGCCGGCTC 1051AAGTGGGCGT GGGTGGAAAA CGGCACGTTC GCAACCGTCG GTAGCGCGCC 1101 GTACCGCGATTTGTCGCCTT TGGGCGCGGA GTGGGCGGAA AAGGCGGATG 1151 GAAATGTCCG CATCGTCGGTTGCGCTGTGT GCGGAGAATT CAAAAAGGCA 1201 CAAGTGCAGG AACAGCTCGC CCGAAAAATCGAGTGGCTGC CGTCTTCCGC 1251 ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAAGAACACGGTT 1301 CCGACCGCTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG CCGCAACGCC1351 TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG CGCTCACCGA 1401TGACGGACAT TATCTCGGGG GAACCATCAT GCCCGGTTTC CACCTGATGA 1451 AAGAATCGCTCGCCGTCCGA ACCGCCAACC TCAACCGGCA CGCCGGTAAG 1501 CGTTATCCTT TCCCGACCACAACGGGCAAT GCCGTCGCCA GCGGCATGAT 1551 GGATGCGGTT TGCGGCTCGG TTATGATGATGCACGGGCGT TTGAAAGAAA 1601 AAACCGGGGC GGGCAAGCCT GTCGATGTCA TCATTACCGGCGGCGGCGCG 1651 GCAAAAGTTG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG AAAATACCGT1701 GCGCGTGGCG GACAACCTCG TCATTTACGG GTTGTTGAAC ATGATTGCCG 1751CCGAAGGCAG GGAATATGAA CATATTTAA

This corresponds to the amino acid sequence <SEQ ID 234; ORF61-1>:

1 MTVLKLSHWR VLAELADGLP QHVSQLARMA DMKPQQLNGF WQQMPAHIRG 51LLRQHDGYWR LVRPLAVFDA EGLRELGERS GFQTALKHEC ASSNDEILEL 101ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS FGWVFDRPQY 151ELGSLSPVAA VACRRALSRL GLDVQIKWPN DLVVGRDKLG GILIETVRTG 201GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA AVLLETLLVE 251LDAVLLQYAR DGFAPFVAEY QAANRDHGKA VLLLRDGETV FEGTVKGVDG 301QGVLHLETAE GKQTVVSGEI SLRSDDRPVS VPKRRDSERF LLLDGGNSRL 351KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KADGNVRIVG CAVCGEFKKA 401QVQEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA LGSRRFSRNA 451CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR TANLNRHAGK 501RYPFPTTTGN AVASGMMDAV CGSVMMMHGR LKEKTGAGKP VDVIITGGGA 551AKVAEALPPA FLAENTVRVA DNLVIYGLLN MIAAEGREYE HI*

FIG. 9 shows plots of hydrophilicity, antigenic index, and AMPHI regionsfor ORF61-1. Further computer analysis of this amino acid sequence gavethe following results:

Homology with the Baf Protein of B. pertussis (Accession Number U12020).

ORF61 and baf protein show 33% aa identity in 166aa overlap:

orf61  23 LLLDGGNSRLKWAWVE-NGTFATVGSAPYR----DLSPLGAEWAEKADGNVRIVGCAVCG  77          +L+D GNSRLK  W + +   A    AP      DL  LG   A      R +G V   Gbaf     3 ILIDSGNSRLKVGWFDPDAPQAAREPAPVAFDNLDLDALGRWLATLPRRPQRALGVNVAG  62orf61  78 EFKKAQVQEQLAR---KIEWLPSSAQAXGIRNHYRHPEEHGSDRW---FNALGSRRFSRN 131             +   +   L      I WL +   A G+RN YR+P++ G+DRW      L  +baf    63 LARGEAIAATLRAGGCDIRWLRAQPLAMGLRNGYRNPDQLGADRWACMVGVLARQPSVHP 122orf61 132 ACVVVSCGTAVTVDALTDDGHYLGXGTIMPGFHLMKESLAVRTANL               177             +V S GTA T+D + D   + G G I+PG  +M+ +LA  TA+Lbaf   123 PLLVASFGTATTLDTIGPDNVFPG-GLILPGPAMMRGALAYGTAHL               167Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF61 shows 97.4% identity over a 189aa overlap with an ORF (ORF61a)from strain A of N. meningitidis:

The complete length ORF61a nucleotide sequence <SEQ ID 235> is:

1 ATGACGGTTT TGAAGCCTTC GCACTGGCGG GTGTTGGCGG AGCTTGCCGA 51CGGTTTGCCG CAACACGTCT CGCAACTGGC GCGTATGGCG GATATGAAGC 101CGCAGCAGCT CAACGGTTTT TGGCAGCAGA TGCCGGCGCA CATACGCGGG 151CTGTTGCGCC AACACGACGG CTATTGGCGG CTGGTGCGCC CATTGGCGGT 201TTTCGATGCC GAAGGTTTGC GCGAGCTGGG GGAAAGGTCG GGTTTTCAGA 251CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT ACTGGAATTG 301GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGTG TGACCCACCT 351GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG CACCGTTTGG 401GCGAGTGTCT GATGTTCAGT TTTGGCTGGG TGTTTGACCG GCCGCAGTAT 451GAGTTGGGTT CGCTGTCGCC TGTTGCGGCA GTGGCGTGCC GGCGCGCCTT 501GTCGCGTTTG GGTTTGAAAA CGCAAATCAA GTGGCCAAAC GATTTGGTCG 551TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACGGT CAGGACGGGC 601GGCAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTCG TGCTGCCCAA 651GGAAGTGGAA AACGCCGCTT CCGTGCAATC GCTGTTTCAG ACGGCATCGC 701GGCGGGGAAA TGCCGATGCC GCCGTGTTGC TGGAAACGCT GTTGGCGGAA 751CTTGATGCGG TGTTGTTGCA ATATGCGCGG GACGGATTTG CGCCTTTTGT 801GGCGGAATAT CAGGCTGCCA ACCGCGACCA CGGCAAGGCG GTATTGCTGT 851TGCGCGACGG CGAAACCGTG TTCGAAGGCA CGGTTAAAGG CGTGGACGGA 901CAAGGCGTTC TGCACTTGGA AACGGCAGAG GGCAAACAGA CGGTCGTCAG 951CGGCGAAATC AGCCTGCGGT CCGACGACAG GCCGGTTTCC GTGCCGAAGC 1001GGCGGGATTC GGAACGTTTT CTGCTGTTGG ACGGCGGCAA CAGCCGGCTC 1051AAGTGGGCGT GGGTGGAAAA CGGCACGTTC GCAACCGTCG GTAGCGCGCC 1101GTACCGCGAT TTGTCGCCTT TGGGCGCGGA GTGGGCGGAA AAGGTGGATG 1151GAAATGTCCG CATCGTCGGT TGCGCCGTGT GCGGAGAATT CAAAAAGGCA 1201CAAGTGCAGG AACAGCTCGC CCGAAAAATC GAGTGGCTGC CGTCTTCCGC 1251ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAA GAACACGGTT 1301CCGACCGCTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG CCGCAACGCC 1351TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG CGCTCACCGA 1401TGACGGACAT TATCTCGGGG GAACCATCAT GCCCGGTTTC CACCTGATGA 1451AAGAATCGCT CGCCGTCCGA ACCGCCAACC TCAACCGGCA CGCCGGTAAG 1501CGTTATCCTT TCCCGACCAC AACGGGCAAT GCCGTCGCCA GCGGCATGAT 1551GGATGCGGTT TGCGGCTCGG TTATGATGAT GCACGGGCGT TTGAAAGAAA 1601AAACCGGGGC GGGCAAGCCT GTCGATGTCA TCATTACCGG CGGCGGCGCG 1651GCAAAAGTTG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG AAAATACCGT 1701GCGCGTGGCG GACAACCTCG TCATTCACGG GCTGCTGAAC CTGATTGCCG 1751CCGAAGGCGG GGAATCGGAA CATACTTAA

This encodes a protein having amino acid sequence <SEQ ID 236>:

1 MTVLKPSHWR VLAELADGLP QHVSQLARMA DMKPQQLNGF WQQMPAHIRG 51LLRQHDGYWR LVRPLAVFDA EGLRELGERS GFQTALKHEC ASSNDEILEL 101ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS FGWVFDRPQY 151ELGSLSPVAA VACRRALSRL GLKTQIKWPN DLVVGRDKLG GILIETVRTG 201GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA AVLLETLLAE 251LDAVLLQYAR DGFAPFVAEY QAANRDHGKA VLLLRDGETV FEGTVKGVDG 301QGVLHLETAE GKQTVVSGEI SLRSDDRPVS VPKRRDSERF LLLDGGNSRL 351KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KVDGNVRIVG CAVCGEFKKA 401QVQEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA LGSRRFSRNA 451CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR TANLNRHAGK 501RYPFPTTTGN AVASGMMDAV CGSVMMMHGR LKEKTGAGKP VDVIITGGGA 551AKVAEALPPA FLAENTVRVA DNLVIHGLLN LIAAEGGESE HT*

ORF61a and ORF61-1 show 98.5% identity in 591 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF61 shows 94.2% identity over a 189aa overlap with a predicted ORF(ORF61.ng) from N. gonorrhoeae:

An ORF61ng nucleotide sequence <SEQ ID 237> was predicted to encode aprotein having amino acid sequence <SEQ ID 238>:

1 MFSFGWAFDR PQYELGSLSP VAALACRRAL GCLGLETQIK WPNDLVVGRD 51KLGGILIETV RAGGKTVAVV GIGINFVLPK EVENAASVQS LFQTASRRGN 101ADAAVLLETL LAELGAVLEQ YAEEGFAPFL NEYETANRDH GKAVLLLRDG 151ETVCEGTVKG VDGRGVLHLE TAEGEQTVVS GEISLRPDNR SVSVPKRPDS 201ERFLLLEGGN SRLKWAWVEN GTFATVGSAP YRDLSPLGAE WAEKADGNVR 251IVGCAVCGES KKAQVKEQLA RKIEWLPSSA QALGIRNHYR HPEEHGSDRW 301FNALGSRRFS RNACVVVSCG TAVTVDALTD DGHYLGGTIM PGFHLMKESL 351AVRTANLNRP AGKRYPFPTT TGNAVASGMM DAVCGSIMMM HGRLKEKNGA 401GKPVDVIITG GGAAKVAEAL PPAFLAENTV RVADNLVIHG LLNLIAAEGG 451 ESEHA*

Further analysis revealed the complete gonococcal DNA sequence <SEQ ID239> to be:

1 ATGACGGTTT TGAAGCCTTC GCATTGGCGG GTGTTGGCGG AGCTTGCCGA 51CGGTTTGCCG CAACACGTAT CGCAATTGGC GCGTGAGGCG GACATGAAGC 101CGCAGCAGCT CAACGGTTTT TGGCAGCAGA TGCCGGCGCA TATACGCGGG 151CTGTTGCGCC AACACGACGG CTATTGGCGG CTGGTGCGCC CCTTGGCGGT 201TTTCGATGCC GAAGGTTTGC GCGATCTGGG GGAAAGGTCG GGTTTTCAGA 251CGGCATTGAA GCACGAGTGC GCGTCCAGCA ACGACGAGAT ACTGGAATTG 301GCGCGGATTG CGCCGGACAA GGCGCACAAA ACCATATGCG TGACCCACCT 351GCAAAGTAAG GGCAGGGGGC GGCAGGGGCG GAAGTGGTCG CACCGTTTGG 401GCGAGTGCCT GATGTTCAGT TTCGGCTGGG CGTTTGACCG GCCGCAGTAT 451GAGTTGGGTT CGCTGTCGCC TGTTGCGGCA CTTGCGTGCC GGCGCGCTTT 501GGGGTGTTTG GGTTTGGAAA CGCAAATCAA GTGGCCAAAC GATTTGGTCG 551TCGGACGCGA CAAATTGGGC GGCATTCTGA TTGAAACAGT CAGGGCGGGC 601GGTAAAACGG TTGCCGTGGT CGGTATCGGC ATCAATTTCG TGCTGCCCAA 651GGAAGTGGAA AACGCCGCTT CCGTGCAGTC GCTGTTTCAG ACGGCATCGC 701GGCGGGGCAA TGCCGATGCC GCCGTATTGC TGGAAACATT GCTTGCGGAA 751CTGGGCGCGG TGTTGGAACA ATATGCGGAA GAAGGGTTCG CGCCATTTTT 801AAATGAGTAT GAAACGGCCA ACCGCGACCA CGGCAAGGCG GTATTGCTGT 851TGCGCGACGG CGAAACCGTG TGCGAAGGCA CGGTTAAAGG CGTGGACGGA 901CGAGGCGTTC TGCACTTGGA AACGGCAgaa ggcgaACAGa cggtcgtcag 951cggcgaaaTC AGcctGCggc ccgacaacaG GTCGGtttcc gtgccgaagc 1001ggccggatTC GgaacgtTTT tTGCtgttgg aaggcgggaa cagccgGCTC 1051AAGTGGGCGT GggtggAAAa cggcacgttc gcaaccgtgg gcagcgcgCc 1101gtaCCGCGAT TTGTCGCCTT TGGGCGCGGA GTGGGCGGAA AAGGCGGATG 1151GAAATGTCCG CATCGTCGGT TGCGCCGTGT GCGGAGAATC CAAAAAGGCA 1201CAAGTGAAGG AACAGCTCGC CCGAAAAATC GAGTGGCTGC CGTCTTCCGC 1251ACAGGCTTTG GGCATACGCA ACCACTACCG CCACCCCGAA GAACACGGTT 1301CCGACCGTTG GTTCAACGCC TTGGGCAGCC GCCGCTTCAG CCGCAACGCC 1351TGCGTCGTCG TCAGTTGCGG CACGGCGGTA ACGGTTGACG CGCTCACCGA 1401TGACGGACAT TATCTCGGCG GAACCATCAT GCCCGGCTTC CACCTGATGA 1451AAGAATCGCT CGCCGTCCGA ACCGCCAACC TCAACCGCCC CGCCGGCAAA 1501CGTTACCCTT TCCCGACCAC AACGGGCAAC GCCGTCGCAA GCGGCATGAT 1551GGACGCGGTT TGCGGCTCGA TAATGATGAT GCACGGCCGT TTGAAAGAAA 1601AAAACGGCGC GGGCAAGCCT GTCGATGTCA TCATTACCGG CGGCGGCGCG 1651GCGAAAGTCG CCGAAGCCCT GCCGCCTGCA TTTTTGGCGG AAAATACCGT 1701GCGCGTGGCG GACAACCTCG TCATCCACGG GCTGCTGAAC CTGATTGCCG 1751CCGAAGGCGG GGAATCGGAA CACGCTTAA

This corresponds to the amino acid sequence <SEQ ID 240; ORF61ng-1>:

1 MTVLKPSHWR VLAELADGLP QHVSQLAREA DMKPQQLNGF WQQMPAHIRG 51LLRQHDGYWR LVRPLAVFDA EGLRDLGERS GFQTALKHEC ASSNDEILEL 101ARIAPDKAHK TICVTHLQSK GRGRQGRKWS HRLGECLMFS FGWAFDRPQY 151ELGSLSPVAA LACRRALGCL GLETQIKWPN DLVVGRDKLG GILIETVRAG 201GKTVAVVGIG INFVLPKEVE NAASVQSLFQ TASRRGNADA AVLLETLLAE 251LGAVLEQYAE EGFAPFLNEY ETANRDHGKA VLLLRDGETV CEGTVKGVDG 301RGVLHLETAE GEQTVVSGEI SLRPDNRSVS VPKRPDSERF LLLEGGNSRL 351KWAWVENGTF ATVGSAPYRD LSPLGAEWAE KADGNVRIVG CAVCGESKKA 401QVKEQLARKI EWLPSSAQAL GIRNHYRHPE EHGSDRWFNA LGSRRFSRNA 451CVVVSCGTAV TVDALTDDGH YLGGTIMPGF HLMKESLAVR TANLNRPAGK 501RYPFPTTTGN AVASGMMDAV CGSIMMMHGR LKEKNGAGKP VDVIITGGGA 551AKVAEALPPA FLAENTVRVA DNLVIHGLLN LIAAEGGESE HA*

ORF61ng-1 and ORF61-1 show 93.9% identity in 591 aa overlap:

Based on this analysis, including the homology with the baf protein ofB. pertussis and the presence of a putative prokaryotic membranelipoprotein lipid attachment site, it is predicted that these proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

Example 29

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 241>:

1 ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT CGTTTATTGC 51CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG GTCGGCGTGC 101GCCTGCTAAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG CCGTCATGTC 151GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG TGTCGTTCGT 201CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG AAATACACTT 251CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT GCTGATGGTG 301TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT ACCACTGGAT 351ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG GCGGGCGGTG 401CGGaAGAGGG CGGCGaAGTC GGCTGGTTCG GCTGCCTGCT GGTGTTGTTG 451GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA GGCTGATTGC 501ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC GCCGCATCGT 551TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA TACCGTGGAC 601TGGAGCGTCG GGATGGTATT GTCGCTGCTG TATTTGGGTT TGGGGTGC..

This corresponds to the amino acid sequence <SEQ ID 242; ORF62>:

1 MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL PALPACRRHV 51GKIPREEWKP LLIVSFVNYV LTLLLQFVGL KYTSAASASV IVGLEPLLMV 101FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV GWFGCLLVLL 151AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS LALAQSYTVD 201WSVGMVLSLL YLGLGC..

Further work revealed the complete nucleotide sequence <SEQ ID 243>:

1 ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT CGTTTATTGC 51CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG GTCGGCGTGC 101GCCTGCTAAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG CCGTCATGTC 151GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG TGTCGTTCGT 201CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG AAATACACTT 251CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT GCTGATGGTG 301TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT ACCACTGGAT 351ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG GCGGGCGGTG 401CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT GGTGTTGTTG 451GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA GGCTGATTGC 501ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC GCCGCATCGT 551TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA TACCGTGGAC 601TGGAGCGTCG GGATGGTATT GTCGCTGCTG TATTTGGGTT TGGGGTGCGG 651CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT GTTCCTGCCA 701ATGTTTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG CGTGCTGCTG 751GCGGTTTTGA TTTTGGGCGA ACACCTGTCG CCCGTGTCCG CCTTGGGCGT 801GTTTGTCGTC ATCGCCGCCA CCTTGGTTGC CGGCCGGCTG TCGCATCAAA 851 AATAA

This corresponds to the amino acid sequence <SEQ ID 244; ORF62-1>:

1 MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL PALPACRRHV 51GKIPREEWKP LLIVSFVNYV LTLLLQFVGL KYTSAASASV IVGLEPLLMV 101FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV GWFGCLLVLL 151AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS LALAQSYTVD 201WSVGMVLSLL YLGLGCGWYA YWLWNKGMSR VPANVSGLLI SLEPVVGVLL 251AVLILGEHLS PVSALGVFVV IAATLVAGRL SHQK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical Transmembrane Protein H10976 of H. influenzae(Accession Number Q57147)

ORF62 and HI0976 show 50% aa identity in 114aa overlap:

Orf62 1 MFYQILALIIWSSSFIAAKYVYGGIDPALMVGVRXXXXXXXXXXXCRRHVGKIPREEWKP 60M YQILAL+IWSSS I  K  Y  +DP L+V VR             R   KI +   K HI0976 1MLYQILALLIWSSSLIVGKLTYSMMDPVLVVQVRLIIAMIIVMPLFLRRWKKIDKPMRKQ 60 Orf62 61LLIVSFVNYVLTLLLQFVGLKYTSAASASVIVGLEPLLMVFVGHFFFNDKARAY 114L  ++F NY    LLQF+GLKYTSA+SA  ++GLEPLL+VFVGHFFF  K   + HI0976 61LWWLAFFNYTAVFLLQFIGLKYTSASSAVTMIGLEPLLVVFVGHFFFKTKQNGF 114Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF62 shows 99.5% identity over a 216aa overlap with an ORF (ORF62a)from strain A of N. meningitidis:

The complete length ORF62a nucleotide sequence <SEQ ID 245> is:

1 ATGTTTTACC AAATCCTTGC CCTGATTATC TGGAGCAGCT CGTTTATTGC 51CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG GTCGGCGTGC 101GCCTGCTGAT TGCTGCGCTG CCTGCACTGC CCGCCTGCCG CCGTCATGTC 151GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG TGTCGTTCGT 201CAACTATGTG CTGACCCTGC TACTTCAGTT TGTCGGGTTG AAATACACTT 251CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCACT GCTGATGGTG 301TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT ACCACTGGAT 351ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG GCGGGCGGTG 401CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT GGTGTTGTTG 451GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA GGCTGATTGC 501ACGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC GCCGCATCGT 551TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA TACCGTGGAC 601TGGAGCGTCG GAATGGTATT GTCGCTGCTG TATTTGGGCG TGGGGTGCAG 651CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT GTTCCTGCCA 701ACGTTTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG CGTGCTGCTG 751GCGGTTTTGA TTTTGGGCGA ACACCTGTCG CCCGTGTCCG TCTTGGGCGT 801GTTTGTCGTC ATCGCCGCCA CCTTGGTTGC CGGCCGGCTG TCGCATCAAA 851 AATAA

This encodes a protein having amino acid sequence <SEQ ID 246>:

1 MFYQILALII WSSSFIAAKY VYGGIDPALM VGVRLLIAAL PALPACRRHV 51GKIPREEWKP LLIVSFVNYV LTLLLQFVGL KYTSAASASV IVGLEPLLMV 101FVGHFFFNDK ARAYHWICGA AAFAGVALLM AGGAEEGGEV GWFGCLLVLL 151AGAGFCAAMR PTQRLIARIG APAFTSVSIA AASLMCLPFS LALAQSYTVD 201WSVGMVLSLL YLGVGCSWYA YWLWNKGMSR VPANVSGLLI SLEPVVGVLL 251AVLILGEHLS PVSVLGVFVV IAATLVAGRL SHQK*

ORF62a and ORF62-1 show 98.9% identity in 284 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF62 shows 99.5% identity over a 216aa overlap with a predicted ORF(ORF62.ng) from N. gonorrhoeae:

The complete length ORF62ng nucleotide sequence <SEQ ID 247> is:

1 ATGTTTTACC AAATCCTTGC CCTGATTATC TGGGGCAGCT CGTTTATTGC 51CGCCAAATAT GTCTATGGCG GCATCGATCC CGCATTGATG GTCGGCGTGC 101GCCTGCTGAT TGCCGCGCTG CCTGCACTGC CCGCCTGCCG CCGTCATGTC 151GGCAAGATTC CGCGTGAGGA ATGGAAGCCG TTGCTGATTG TGTCGTTCGT 201CAACTATGTG CTGACCCTGC TGCTTCAGTT TGTCGGGTTG AAATACACTT 251CCGCCGCCAG CGCATCGGTC ATTGTCGGAC TCGAGCCGCT GCTGATGGTG 301TTTGTCGGAC ACTTTTTCTT CAACGACAAA GCGCGTGCCT ACCACTGGAT 351ATGCGGCGCG GCGGCATTTG CCGGTGTCGC GCTGCTGATG GCGGGCGGTG 401CGGAAGAGGG CGGCGAAGTC GGCTGGTTCG GCTGCCTGCT GGTGTTGTTG 451GCGGGCGCGG GCTTTTGTGC CGCTATGCGT CCGACGCAAA GGCTGATTGC 501CCGCATCGGC GCACCGGCAT TCACATCTGT TTCCATTGCC GCCGCATCGT 551TGATGTGCCT GCCGTTTTCG CTTGCTTTGG CGCAAAGTTA TACCGTGGAC 601TGGAGCGTCG GGATGGTATT GTCGCTGTTG TATTTGGGTT TGGGGTGCGG 651CTGGTACGCC TATTGGCTGT GGAACAAGGG GATGAGCCGT GTTCCTGCCA 701ACGCGTCGGG ACTGTTGATT TCGCTCGAAC CCGTCGTCGG CGTGCTGTTG 751GCGGTTTTGA TTTTGGGCGA ACATTTATCG CCCGTGTCCG CCTTGGGCGT 801GTTTGTCGTC ATCGCCGCCA CTTTCGCCGC CGGCCGGCTG TCGCGCAGGG 851ACGCGCAAAA CGGCAATGCC GTCTGA

This encodes a protein having amino acid sequence <SEQ ID 248>:

1 MFYQILALII WGSSFIAAKY VYGGIDPALM VGVRLLIAAL PALPACRRHV 51 GKIPREEWKPLLIVSFVNYV LTLLLQFVGL KYTSAASASV IVGLEPLLMV 101 FVGHFFFNDKARAYHWICGA AAFAGVALLM AGGAEEGGEV GWFGCLLVLL 151 AGAGFCAAMR PTQRLIARIGAPAFTSVSIA AASLMCLPFS LALAQSYTVD 201 WSVGMVLSLL YLGLGCGWYA YWLWNKGMSRVPANASGLLI SLEPVVGVLL 251 AVLILGEHLS PVSALGVFVV IAATFAAGRL SRRDAQNGNA V*

ORF62ng and ORF62-1 show 97.9% identity in 283 aa overlap:

Furthermore, ORF62ng shows significant homology to a hypothetical H.influenzae protein:

sp|Q57147|Y976_HAEIN HYPOTHETICAL PROTEIN HI0976 >gi|1074589|pir||B64163hypothetical protein HI0976 - Haemophilus influenzae (strain RdKW20) >gi|1574004 (U32778) hypothetical [Haemophilus influenzae] Length= 128 Score = 106 bits (262), Expect = 2e−22 Identities = 56/114 (49%),Positives = 68/114 (59%) Query: 1MFYQILALIIWGSSFIAAKYVYGGIDPALMVGVRXXXXXXXXXXXCRRHVGKIPREEWKP 60 MYQILAL+IW SS I  K  Y  +DP L+V VR             R   KI +   K Sbjct: 1MLYQILALLIWSSSLIVGKLTYSMMDPVLVVQVRLIIAMIIVMPLFLRRWKKIDKPMRKQ 60 Query:61 LLIVSFVNYVLTLLLQFVGLKYTSAASASVIVGLEPLLMVFVGHFFFNDKARAY 114 L  ++FNY    LLQF+GLKYTSA+SA  ++GLEPLL+VFVGHFFF  K   + Sbjct: 61LWWLAFFNYTAVFLLQFIGLKYTSASSAVTMIGLEPLLVVFVGHFFFKTKQNGF 114

Based on this analysis, including the homology with the transmembraneprotein of H. influenzae and the putative leader sequence and severaltransmembrane domains in the gonococcal protein, it is predicted thatthese proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 30

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 249>:

1 ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCmGwms TCCTGkkGTA 51 sGGACTGACGGCGGCAACCG GCAGCACCAG TTCGCTGGCG GATTATTTCT 101 GGTGGATTGT TGCGTTCAGCGCAATGCTGC TGCTGGTGTT GTCCGCCGTT 151 TTGGCACGTT ATGTCATATT GCTGTTGAAAGACAGGCGCG ACGGCGTATT 201 CGGTTCGCtA srTyGCCAAA gsGCCTgkks TGGG.ATGTTTACGCTGGTT 251 GCCGkACTGC CCGGCGTGTT TCTGTTCGGC TTTCCCGCAC AGTTCATCAA301 CGGCACGATT AATTCGTGGT TCGGCAACGA TACCCACGAG GCGCTTGAAC 351GCAGCCTCAA TTTGAGCAAG TCCGCATTGA ATTTGGCGGC AGACAACGCC 401 CTCGGCAACGCCGTCCCCGT GCAGATAGAC CTCATCGGCG CGGCTTCCCT 451 GCCCGGGGAT ATGGGCAGGGTGCTGGAACA TTACGCCGGC AGCGGTTTTG 501 CCCAGCTTGC CCTGTACAAy ksCGCAAGCGGCAAAATCGA AAAAAGCATC 551 AACCCGCACA AGCTCGATCA GCCGTTTCCA GGTAAGGCGCGTTGGGAaAa 601 AATCCaACGG GCGGGTTCGG TCAGGGATTT GGAAAGCATA GGCGGCGTAT651 TGTaCGCGCA GGGCTGGCTG TCGGCGGGTA CGCACwACGG GCGCGATTAC 701GCCTTGTTTT TCCGTCAGCC GGTTCCCAAA GGCGTGGCAG AGGATGCCGT 751 yTTAATCGAAAAGGCAAGGG CGAAATATGC TGAGTTGAGT TACAGCAAAA 801 AAGGTTTGCA GACCTTTTTCCTGGCAACCC TGCTGATTGC CTCGCTGCTG 851 TCGATTTTTC TTGCACTGGT CATGGCACTGTATTTCGCCC GCCGTTTCGT 901 CGAACCCGTC CTATCGCTTG CCGAGGGGGC GAAGGCGGTGGCGCAAGGCG 951 ATTTCAGCCA GACGCGCCCC GTGTTGCGCA ACGACGAGTT CGGACGCTTG1001 ACCArGTTGT TCAACCACAT GACCGAGCAG CTTTCCATCG CCAAAGATGC 1051AGACGAGCGC AACCGCCGGC GCGAGGAAGC CGCCAGGCAT TATCTTGAAT 1101 GCGTGTTGGAGGGGCTGACC ACGGGCGTGG TGGTGTTTGA CGAACAAGGC 1151 TGTCTGAAAA CCTTCAACAAAGCGGCGGGT ACC..

This corresponds to the amino acid sequence <SEQ ID 250; ORF64>:

1 MRRFLPIAAI CAXXLXXGLT AATGSTSSLA DYFWWIVAFS AMLLLVLSAV 51 LARYVILLLKDRRDGVFGSX XAKXPXXXMF TLVAXLPGVF LFGFPAQFIN 101 GTINSWFGND THEALERSLNLSKSALNLAA DNALGNAVPV QIDLIGAASL 151 PGDMGRVLEH YAGSGFAQLA LYNXASGKIEKSINPHKLDQ PFPGKARWEK 201 IQRAGSVRDL ESIGGVLYAQ GWLSAGTHXG RDYALFFRQPVPKGVAEDAV 251 LIEKARAKYA ELSYSKKGLQ TFFLATLLIA SLLSIFLALV MALYFARRFV301 EPVLSLAEGA KAVAQGDFSQ TRPVLRNDEF GRLTXLFNHM TEQLSIAKDA 351DERNRRREEA ARHYLECVLE GLTTGVVVFD EQGCLKTFNK AAGT..

Further work revealed the complete nucleotide sequence <SEQ ID 251>:

1 ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCCGTCG TCCTGTTGTA 51 CGGACTGACGGCGGCAACCG GCAGCACCAG TTCGCTGGCG GATTATTTCT 101 GGTGGATTGT TGCGTTCAGCGCAATGCTGC TGCTGGTGTT GTCCGCCGTT 151 TTGGCACGTT ATGTCATATT GCTGTTGAAAGACAGGCGCG ACGGCGTATT 201 CGGTTCGCAG ATTGCCAAAC GCCTTTCTGG GATGTTTACGCTGGTTGCCG 251 TACTGCCCGG CGTGTTTCTG TTCGGCGTTT CCGCACAGTT CATCAACGGC301 ACGATTAATT CGTGGTTCGG CAACGATACC CACGAGGCGC TTGAACGCAG 351CCTCAATTTG AGCAAGTCCG CATTGAATTT GGCGGCAGAC AACGCCCTCG 401 GCAACGCCGTCCCCGTGCAG ATAGACCTCA TCGGCGCGGC TTCCCTGCCC 451 GGGGATATGG GCAGGGTGCTGGAACATTAC GCCGGCAGCG GTTTTGCCCA 501 GCTTGCCCTG TACAATGCCG CAAGCGGCAAAATCGAAAAA AGCATCAACC 551 CGCACAAGCT CGATCAGCCG TTTCCAGGTA AGGCGCGTTGGGAAAAAATC 601 CAACGGGCGG GTTCGGTCAG GGATTTGGAA AGCATAGGCG GCGTATTGTA651 CGCGCAGGGC TGGCTGTCGG CGGGTACGCA CAACGGGCGC GATTACGCCT 701TGTTTTTCCG TCAGCCGGTT CCCAAAGGCG TGGCAGAGGA TGCCGTCTTA 751 ATCGAAAAGGCAAGGGCGAA ATATGCTGAG TTGAGTTACA GCAAAAAAGG 801 TTTGCAGACC TTTTTCCTGGCAACCCTGCT GATTGCCTCG CTGCTGTCGA 851 TTTTTCTTGC ACTGGTCATG GCACTGTATTTCGCCCGCCG TTTCGTCGAA 901 CCCGTCCTAT CGCTTGCCGA GGGGGCGAAG GCGGTGGCGCAAGGCGATTT 951 CAGCCAGACG CGCCCCGTGT TGCGCAACGA CGAGTTCGGA CGCTTGACCA1001 AGTTGTTCAA CCACATGACC GAGCAGCTTT CCATCGCCAA AGAAGCAGAC 1051GAGCGCAACC GCCGGCGCGA GGAAGCCGCC AGGCATTATC TTGAATGCGT 1101 GTTGGAGGGGCTGACCACGG GCGTGGTGGT GTTTGACGAA CAAGGCTGTC 1151 TGAAAACCTT CAACAAAGCGGCGGAACAGA TTTTGGGGAT GCCGCTTACC 1201 CCCCTGTGGG GCAGCAGCCG GCACGGTTGGCACGGCGTTT CGGCGCAGCA 1251 GTCCCTGCTT GCCGAAGTGT TTGCCGCCAT CGGCGCGGCGGCAGGTACGG 1301 ACAAACCGGT CCATGTGAAA TATGCCGCGC CGGACGATGC CAAAATCCTG1351 CTGGGCAAGG CAACCGTCCT GCCCGAAGAC AACGGCAACG GCGTGGTAAT 1401GGTGATTGAC GACATCACCG TTTTGATACA CGCGCAAAAA GAAGCCGCGT 1451 GGGGCGAAGTGGCGAAGCGG CTGGCACACG AAATCCGCAA TCCGCTCACG 1501 CCCATCCAGC TTTCCGCCGAACGGCTGGCG TGGAAATTGG GCGGGAAGCT 1551 GGATGAGCAG GATGCGCAAA TCCTGACGCGTTCGACCGAC ACCATCGTCA 1601 AACAGGTGGC GGCATTGAAG GAAATGGTCG AAGCATTCCGCAATTATGCG 1651 CGTTCCCCTT CGCTCAAATT GGAAAATCAG GATTTGAACG CCTTAATCGG1701 CGATGTGTTG GCATTGTATG AAGCCGGTCC GTGCCGGTTT GCGGCGGAGC 1751TTGCCGGCGA ACCGCTGACG GTGGCGGCGG ATACGACCGC CATGCGGCAG 1801 GTGCTGCACAATATTTTCAA AAATGCCGCC GAAGCGGCGG AAGAAGCCGA 1851 TGTGCCCGAA GTCAGGGTAAAATCGGAAAC AGGGCAGGAC GGTCGGATTG 1901 TCCTGACGGT TTGCGACAAC GGCAAAGGGTTCGGCAGGGA AATGCTGCAC 1951 AACGCCTTCG AGCCGTATGT AACGGACAAA CCGGCGGGAACGGGATTGGG 2001 TCTGCCTGTG GTGAAAAAAA TCATTGAAGA ACACGGCGGC CGCATCAGCC2051 TGAGCAATCA GGATGCGGGT GGCGCGTGTG TCAGAATCAT CTTGCCAAAA 2101ACGGTAAAAA CTTATGCGTA G

This corresponds to the amino acid sequence <SEQ ID 252; ORF64-1>:

1 MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVAFS AMLLLVLSAV 51 LARYVILLLKDRRDGVFGSQ IAKRLSGMFT LVAVLPGVFL FGVSAQFING 101 TINSWFGNDT HEALERSLNLSKSALNLAAD NALGNAVPVQ IDLIGAASLP 151 GDMGRVLEHY AGSGFAQLAL YNAASGKIEKSINPHKLDQP FPGKARWEKI 201 QRAGSVRDLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPVPKGVAEDAVL 251 IEKARAKYAE LSYSKKGLQT FFLATLLIAS LLSIFLALVM ALYFARRFVE301 PVLSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT EQLSIAKEAD 351ERNRRREEAA RHYLECVLEG LTTGVVVFDE QGCLKTFNKA AEQILGMPLT 401 PLWGSSRHGWHGVSAQQSLL AEVFAAIGAA AGTDKPVHVK YAAPDDAKIL 451 LGKATVLPED NGNGVVMVIDDITVLIHAQK EAAWGEVAKR LAHEIRNPLT 501 PIQLSAERLA WKLGGKLDEQ DAQILTRSTDTIVKQVAALK EMVEAFRNYA 551 RSPSLKLENQ DLNALIGDVL ALYEAGPCRF AAELAGEPLTVAADTTAMRQ 601 VLHNIFKNAA EAAEEADVPE VRVKSETGQD GRIVLTVCDN GKGFGREMLH651 NAFEPYVTDK PAGTGLGLPV VKKIIEEHGG RISLSNQDAG GACVRIILPK 701 TVKTYA*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF64 shows 92.6% identity over a 392aa overlap with an ORF (ORF64a)from strain A of N. meningitidis:

The complete length ORF64a nucleotide sequence <SEQ ID 253> is:

1 ATGCGCCGTT TTCTACCGAT CGCAGCCATA TGCGCCGTCG TCCTGTTGTA 51 CGGACTGACGGCGGCAACCG GCAGCACCAG TTCGCTGGCG GATTATTTCT 101 GGTGGATTGT TGCGTTCAGCGCAATGCTGC TGCTGGTGTT GTCCGCCGTT 151 TTGGCACGTT ATGTCATATT GCTGTTGAAAGACAGGCGCG ACGGCGTATT 201 CGGTTCGCAG ATTGCCAAAC GCCTTTCCGG GATGTTTACGCTGGTTGCCG 251 TACTGCCCGG CGTGTTTCTG TTCGGCGTTT CCGCACAGTT TATCAACGGC301 ACGATTAATT CGTGGTTCGG CAACGATACC CACGAGGCGC TTGAACGCAG 351CCTCAATTTG AGCAAGTCCG CATTGAATCT GGCGGCAGAC AACGCCCTTG 401 GCAACGCCATCCCCGTGCAG ATAGACNTCA TCGGCGCGGC TTCCCTGCCC 451 NGGGATATGG GCAGGGTGCTGGAACATTAC GCCGGCAGCG GTTTTGCCCA 501 GCTTGCCCTG TACAATGCCG CAAGCGGCAAAATCGAAAAA AGCATCAACC 551 CGCACAAGCT CGATCAGCCG TTTCCAGGTA AGGCGCGTTGGGAAAAAATC 601 CAACAGGCGG GTTCGGTCAG GGATNNGGAA AGCATAGGCG GCGTATTGTA651 CGCGCANGGC TGGCTGTCGG CAGNNACGCA CAACGGGCGC GATTACGCCT 701TGTTTTTCCG TCAGCCGGTT CCCAAAGGCG TGGCAGAGGA TGCCGTCTTA 751 ATCGAAAAGGCAAGGGCGNA ANANNNTNAG TTGAGTTACA GCAAAAAAGG 801 TTTGCAGACC TTTTTCCTNGCAACCCTGCT GATTGCCTCN CTGCTGTCGA 851 TTTTTCTTGC ACTGGTCATG GCACTGTATTTCGCCCGCCG TTTCGTCGAA 901 CCCGTCCTAT CGCTTGCCGA GGGGGCGAAG GCGGTGGCGCAAGGCGATTT 951 CAGCCAGACG CGCCCCGTGT TGCGCAACGA CGAGTTCGGA CGCTTGACCA1001 AGTTGTTCAA CCACATGACC GAGCAGCTTT CCATCGCCAA AGAAGCAGAC 1051GAGCGCAACC GCCGGCGCGA GGAAGCCGCC AGACATTATC TCGAATGCGT 1101 GTTGGAGGGGCTGACCACGG GCGTGGTGGT GTTTGACGAA CAAGGCTGTC 1151 TGAAAACCTT CAACAAAGCGGCGGAACAGA TTTTGGGGAT GCCGCTTACC 1201 CCCCTGTGGG GCAGCAGCCG GCACGGTTGGCACGGCGTTT CGGCGCAGCA 1251 GTCCCTGCTT GCCGAAGTGT TTGCCGCCAT CGGCGCGGCGGCAGGTACGG 1301 ACAAACCGGT CCATGTGAAA TATGCCGCGC CGGACGATGC CAAAATCCTG1351 CTGGGCAAGG CAACCGTCCT GCCCGAAGAC AACNGCAACG GCGTGGTAAT 1401GGTGATTGAC GACATCACCG TTTTGATACA CGCGCAAAAA GAAGCCGCGT 1451 GGGGCGAAGTGGCAAAACGG CTGGCACACG AAATCCGCAA TCCGCTCACG 1501 CCCATCCAGC TTTCTGCCGAACGGCTGGCG TGGAAATTGG GCGGGAAGCT 1551 GGACGAGCAN GACGCGCAAA TCCTGACACGTTCGACCGAC ACCATCATCA 1601 AACAAGTGGC GGCATTAAAA GAAATGGTCG AGGCATTCCGCAATTACNCG 1651 CGTTCCCCTT CGNCTCAATT GGAAAATCAG GATTTGAACG CCTTAATCGG1701 CGATGTGTTG GCATTGTACG AAGCTGGTCC GTGCCGGTTT GCGGCGGAAC 1751TTGCCGGCGA ACCGCTGATG ATGGCGGCGG ATACGACCGC CATGCGGCAG 1801 GTGCTGCACAATATTTTCAA AAATGCCGCC GAAGCGGCGG AAGAAGCCGA 1851 TGTGCCCGAA GTCAGGGTAAAATCGGAAGC GGGGCAGGAC GGACGGATTG 1901 TCCTGACAGT TTGCGACAAC GGCAAGGGGTTCGGCAGGGA AATGCTGCAC 1951 AATGCCTTCG AGCCGTATGT AACGGACAAA CCGGCTGGAACGGGATTGNG 2001 ACTGCCCGTG GTGAAAAAAA TCATTGAAGA ACACGGCGGC CNCATCAGCC2051 TGAGCAATCA GGATGCGGGC GGCGCGTNTG TCAGAATCAT CTTGCCAAAA 2101ACGGTAGAAA CTTATGCGTA G

This encodes a protein having amino acid sequence <SEQ ID 254>:

1 MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVAFS AMLLLVLSAV 51 LARYVILLLKDRRDGVFGSQ IAKRLSGMFT LVAVLPGVFL FGVSAQFING 101 TINSWFGNDT HEALERSLNLSKSALNLAAD NALGNAIPVQ IDXIGAASLP 151 XDMGRVLEHY AGSGFAQLAL YNAASGKIEKSINPHKLDQP FPGKARWEKI 201 QQAGSVRDXE SIGGVLYAXG WLSAXTHNGR DYALFFRQPVPKGVAEDAVL 251 IEKARAXXXX LSYSKKGLQT FFLATLLIAS LLSIFLALVM ALYFARRFVE301 PVLSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT EQLSIAKEAD 351ERNRRREEAA RHYLECVLEG LTTGVVVFDE QGCLKTFNKA AEQILGMPLT 401 PLWGSSRHGWHGVSAQQSLL AEVFAAIGAA AGTDKPVHVK YAAPDDAKIL 451 LGKATVLPED NXNGVVMVIDDITVLIHAQK EAAWGEVAKR LAHEIRNPLT 501 PIQLSAERLA WKLGGKLDEX DAQILTRSTDTIIKQVAALK EMVEAFRNYX 551 RSPSXQLENQ DLNALIGDVL ALYEAGPCRF AAELAGEPLMMAADTTAMRQ 601 VLHNIFKNAA EAAEEADVPE VRVKSEAGQD GRIVLTVCDN GKGFGREMLH651 NAFEPYVTDK PAGTGLXLPV VKKIIEEHGG XISLSNQDAG GAXVRIILPK 701 TVETYA*

ORF64a and ORF64-1 show 96.6% identity in 706 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF64 shows 86.6% identity over a 387aa overlap with a predicted ORF(ORF64.ng) from N. gonorrhoeae:

An ORF64ng nucleotide sequence <SEQ ID 255> was predicted to encode aprotein having amino acid sequence <SEQ ID 256>:

1 MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVSFS AMLLLVLSAV 51 LARYVILLLKDRRNGVFGSQ IAKRLSGMFT LVAVLPGLFL FGISAQFING 101 TINSWFGNDT HEALERSLNLSKSALDLAAD NAVSNAVPVQ IDLIGTASLS 151 GNMGSVLEHY AGSGFAQLAL YNAASGKIEKSINPHQFDQP LPDKEHWEQI 201 QQTGSVRSLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPIPENVAQDAVL 251 IEKARAKYAE LSYSKKGLQT FFLVTLLIAS LLSIFLALVM ALYFARRFVE301 PILSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT EQLSIAKEAD 351ERNRRREEAA RHYLECVLDG LTTGVVVSYP LSCCRTAVFS TCHSSPLSYF*

Further work revealed the complete gonococcal DNA sequence <SEQ ID 257>:

1 ATGCGCCGCT TCCTACCGAT CGCAGCCATA TGCGCCGTCG TCCTGCTGTA 51 CGGATTGACGGCGGCGACCG GCAGCACCAG TTCGCTGGCG GATTATTTCT 101 GGTGGATAGT CTCGTTCAGCGCAATGCTGC TGCTGGTGTT GTCCGCCGTT 151 TTGGCACGTT ATGTCATATT GCTGTTGAAAGACAGGCGCA ACGGCGTGTT 201 CGGTTCGCAG ATTGCCAAAC GCCTTTCCGG GATGTTCACGCTGGTCGCCG 251 TACTGCCCGG CTTGTTCCTG TTCGGCATTT CCGCGCAGTT TATCAACGGC301 ACGATTAATT CGTGGTTCGG CAACGACACC CACGAAGCCC TCGAACGCAG 351CCTTAATTTG AGCAAGTCCG CACTGGATTT GGCGGCAGAC AATGCCGTCA 401 GCAACGCCGTTCCCGTACAG ATAGACCTCA TCGGCACCGC CTCCCTGTCG 451 GGCAATATGG GCAGTGTGCTGGAACACTAC GCCGGCAGCG GTTTTGCCCA 501 GCTTGCCCTG TACAATGCCG CAAGCGGGAAAATCGAAAAA AGCATCAATC 551 CGCACCAATT CGACCAGCCG CTTCCCGACA AAGAACATTGGGAACAGATT 601 CAGCAGACCG GTTCGGTTCG GAGTTTGGAA AGCATAGGCG GCGTATTGTA651 CGCGCAGGGA TGGTTGTCGG CAGGTACGCA CAACGGGCGC GATTACGCGC 701TGTTCTTCCG CCAGCCGATT CCCGAAAATG TGGCACAGGA TGCCGTTCTG 751 ATTGAAAAGGCGCGGGCGAA ATATGCCGAA TTGAGTTACA GCAAAAAAGG 801 TTTGCAGACC TTTTTTCTGGTAACCCTGCT GATTGCCTCG CTGCTGTCGA 851 TTTTTCTTGC GCTGGTAATG GCACTGTATTTTGCCCGCCG TTTCGTCGAA 901 CCCATTCTGT CGCTTGCCGA GGGCGCAAAG GCGGTGGCGCAGGGTGATTT 951 CAGCCAGACG CGCCCCGTAT TGCGCAACGA CGAGTTCGGA CGTTTGACCA1001 AGCTGTTCAA CCATATGACC GAGCAGCTTT CCATCGCCAA AGAAGCAGAC 1051GAACGCAACC GCCGGCGCGA GGAAGCCGCC CGTCACTACC TCGAGTGCGT 1101 GTTGGATGGGTTGACTACCG GTGTGGTGGT GTTTGACGAA AAAGGCCGTT 1151 TGAAAACCTT CAACAAGGCGGCGGAACAGA TTTTGGGGAT GCCGCTCGCC 1201 CCCCTGTGGG GCAGCAGCCG GCACGGTTGGCACGGCGTTT CGGCGCAGCA 1251 GTCCCTGCTT GCCGAAGTGT TtgccgccAT CGGTGCGGCGGCAGGTACGG 1301 ACAAACCGGT CCAGGTGGAA TATGCCGCGC CGGACGATGC CAAAATCCTG1351 CTGGGCAAGG CGACGGTATT GCCCGAAGAC AACGGCAACG GCGTGGTGAT 1401GGTGATTGAC GACATCACCG TGCTGATACG CGCGCAAAAA GAAGCCGCGT 1451 GGGGTGAAGTGGCGAAGCGG CTGGCACACG AAATCCGCAA TCCGCTCACG 1501 CCCATCCAGC TTTCCGCCGAACGGCTGGCG TGGAAATTGG GCGGGAAGCT 1551 GGACGATCAG GACGCGCAAA TCCTGACGCGTtcgACCGAC ACCATCATCA 1601 AACAGgtggc gGCGTTAAAA GAAATGGTCG AGGCATTCCGCAATTACGCG 1651 CGCGCCCCTT CGCTCAAACT GGAAAATCAG GATTTGAACG CCTTAATCGG1701 CGATGTTTTG GCCCTGTACG AAGCCGGCCC GTGCCGGTTT GAGGCGGAAC 1751TTGCCGGCGA ACCGCTGATG ATGGCGGCGG ATACGACCGC CATGCGGCAG 1801 GTGCTGCACAATATTTTCAA AAATGCCGCC GAAGCGGCGG AAGAAGCCGA 1851 TATGCCCGAA GTCAGGGTAAAATCGGAAAC GGGGCAGGAC GGACGGATTG 1901 TCCTGACGGT TTGCGACAAC GGCAAGGGATTCGGCAAGGA AATGCTGCAC 1951 AATGCTTTCG AGCCGTATGT GACGGATAAG CCGGCGGGAACGGGACTGGG 2001 TCTGCCTGTA GTGAAAAAAA TCATTGGAGA ACACGGCGGC CGCATCAGCC2051 TGAGCAATCA GGATGCGGGT GGGGCGTGTG TCAGAATCAT CTTGCCAAAA 2101ACGGTAGAAA CTTATGCGTA G

This corresponds to the amino acid sequence <SEQ ID 258; ORF64ng-1>:

1 MRRFLPIAAI CAVVLLYGLT AATGSTSSLA DYFWWIVSFS AMLLLVLSAV 51 LARYVILLLKDRRNGVFGSQ IAKRLSGMFT LVAVLPGLFL FGISAQFING 101 TINSWFGNDT HEALERSLNLSKSALDLAAD NAVSNAVPVQ IDLIGTASLS 151 GNMGSVLEHY AGSGFAQLAL YNAASGKIEKSINPHQFDQP LPDKEHWEQI 201 QQTGSVRSLE SIGGVLYAQG WLSAGTHNGR DYALFFRQPIPENVAQDAVL 251 IEKARAKYAE LSYSKKGLQT FFLVTLLIAS LLSIFLALVM ALYFARRFVE301 PILSLAEGAK AVAQGDFSQT RPVLRNDEFG RLTKLFNHMT EQLSIAKEAD 351ERNRRREEAA RHYLECVLDG LTTGVVVFDE KGRLKTFNKA AEQILGMPLA 401 PLWGSSRHGWHGVSAQQSLL AEVFAAIGAA AGTDKPVQVE YAAPDDAKIL 451 LGKATVLPED NGNGVVMVIDDITVLIRAQK EAAWGEVAKR LAHEIRNPLT 501 PIQLSAERLA WKLGGKLDDQ DAQILTRSTDTIIKQVAALK EMVEAFRNYA 551 RAPSLKLENQ DLNALIGDVL ALYEAGPCRF EAELAGEPLMMAADTTAMRQ 601 VLHNIFKNAA EAAEEADMPE VRVKSETGQD GRIVLTVCDN GKGFGKEMLH651 NAFEPYVTDK PAGTGLGLPV VKKIIGEHGG RISLSNQDAG GACVRIILPK 701 TVETYA*

ORF64ng-1 and ORF64-1 show 93.8% identity in 706 aa overlap:

Furthermore, ORF64ng-1 shows significant homology to a protein from A.caulinodans:

sp|Q04850|NTRY_AZOCA NITROGEN REGULATION PROTEINNTRY >gi|77479|pir||S18624 ntrY protein - Azorhizobiumcaulinodans >gi|38737 (X63841) NtrY gene product [Azorhizobiumcaulinodans] Length = 771 Score = 218 bits (550), Expect = 7e−56Identities = 195/720 (27%), Positives = 320/720 (44%), Gaps = 58/720(8%) Query: 7IAAICAVVLLYGLTAATGSTSSLADYFWWIXXXXXXXXXXXXXXXXRYVILLLKDRRNGV 66I+A+   ++L GLT    +   +      +                R +  + K R  G Sbjct: 35ISALATFLILMGLTPVVPTHQVVIS----VLLVNAAAVLILSAMVGREIWRIAKARARGR 90 Query:67 FGSQIAKRLSGMFTLVAVLPGLFLFGISAQFINGTINSWFGNDTHEALERSLNLSKSALD 126  +++  R+ G+F +V+V+P + +  +++  ++  ++ WF   T E +  S++++++ + Sbjct: 91AAARLHIRIVGLFAVVSVVPAILVAVVASLTLDRGLDRWFSMRTQEIVASSVSVAQTYVR 150 Query:127 LAADNAVSNAVPVQIDLIGTASLSGNMGSVLEHYAG--SGFAQLALYNAASGKIEKSINP 184   AN   + + +  DL    S+          Y G  S F Q+    AA   +  ++ Sbjct: 151EHALNIRGDILAMSADLTRLKSV----------YEGDRSRFNQILTAQAALRNLPGAMLI 200 Query:185 HQFDQPLPDKEHWEQIQQTGSVRSLESIGGVLYAQGWLSAGTHNGRDYA----------- 233 + D  + ++ +   I +   V +  +IG     Q  +     N  DY Sbjct: 201RR-DLSVVERAN-VNIGREFIVPANLAIGDATPDQPVIYLP--NDADYVAAVVPLKDYDD 256 Query:234 --LFFRQPIPENVAQDAVLIEKARAKYAELSYSKKGLQTFFLVTXXXXXXXXXXXXXVMA 291  L+  + I   V       ++  A Y  L   + G+Q  F +               + Sbjct: 257LYLYVARLIDPRVIGYLKTTQETLADYRSLEERRFGVQVAFALMYAVITLIVLLSAVWLG 316 Query:292 LYFARRFVEPILSLAEGAKAVAQGDFSQTRPVLRND-EFGRLTKLFNHMTEQLSIXXXXX 350 LF++  V PI  L   A  VA+G+     P+ R + +   L + FN MT +L Sbjct: 317LNFSKWLVAPIRRLMSAADHVAEGNLDVRVPIYRAEGDLASLAETFNKMTHELRSQREAI 376 Query:351 XXXXXXXXXXXHYLECVLDGLTTGVVVFDEKGRLKTFNKAAEQILGMPLAPLWGSSRHGW 410            + E VL G+  GV+  D + R+   N++AE++LG  L+ +    RH Sbjct: 377LTARDQIDSRRRFTEAVLSGVGAGVIGLDSQERITILNRSAERLLG--LSEVEALHRHLA 434 Query:411 HGVSAQQSLLAEVFXXXXXXXXTDKPVQVEYAAPDDAKILLGKATVLPEDNG---NGVVM 467  V     LL E            + VQ       D +  +    V  E +    +G V+ Sbjct: 435EVVPETAGLLEEA------EHARQRSVQGNITLTRDGRERVFAVRVTTEQSPEAEHGWVV 488 Query:468 VIDDITVLIRAQKEAAWGEVAKRLAHEIRNPLTPIQLSAERLAWKLGGKLDDQDAQILTR 527 +DDIT LI AQ+ +AW +VA+R+AHEI+NPLTPIQLSAERL  K G  +  QD +I  + Sbjct: 489TLDDITELISAQRTSAWADVARRIAHEIKNPLTPIQLSAERLKRKFGRHV-TQDREIFDQ 547 Query:528 STDTIIKQVAALKEMVEAFRNYARAPSLKLENQDLNALIGDVLALYEAGPCRFEAELAGE 587 TDTII+QV  +  MV+ F ++AR P   +++QD++ +I   + L   G      + Sbjct: 548CTDTIIRQVGDIGRMVDEFSSFARMPKPVVDSQDMSEIIRQTVFLMRVGHPEVVFDSEVP 607 Query:588 PLMMAA-DTTAMRQVLHNIFKNXXXXXXXXDMPEVRVK-------SETGQDGRIVLTVCD 639 P MA  D   + Q L NI KN          P+VR +       +  G+D  +V+ + D Sbjct: 608PAMPARFDRRLVSQALTNILKNAAEAIEAVP-PDVRGQGRIRVSANRVGED--LVIDIID 664 Query:640 NGKGFGKEMLHNAFEPYVTDKPAGTGLGLPVVKKIIGEHGGRISLSNQDAG-GACVRIIL 698 NGG  +E  +   EPYVT +  GTGLGL +V KI+ EHGG I L++   G GA +R+ L Sbjct: 665NGTGLPQESRNRLLEPYVTTREKGTGLGLAIVGKIMEEHGGGIELNDAPEGRGAWIRLTL 724

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 31

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 259>:

1 ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT TCCGGCTGGT 51 GCTTTTTCATATCCTCATCA TCGCCGCCAG CAACTATCTG GTGCAGTTCC 101 CTTTCCAAAT TTTCGGCATCCACACCACTT GGGGCGCATT TTCCTTTCCC 151 TTCATCTTCC TTGCCACCGA CCTGACCGTCCGCATTTTCG GTTCTCACTT 201 GGCACGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTTTTGCTTTCCT 251 ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACAGG CTTGGGCGCG301 CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCCTTAG CCAGCTTTGC 351CGCCTACGCG ATCGGACAAA TCCTTGATAT TTTTGTATTC AACAAATTAC 401 GCCGTCTGAAAGCGTGGTGG ATTGCACCGA ACGCATCAAC CGTCATCGGG 451 CACGCGTTGG ATACG...

This corresponds to the amino acid sequence <SEQ ID 260; ORF66>:

1 MYAFTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFQIFGI HTTWGAFSFP 51 FIFLATDLTVRIFGSHLARR IIFWVMFPAL LLSYVFSVLF HNGSWTGLGA 101 LSEFNTFVGR IALASFAAYAIGQILDIFVF NKLRRLKAWW IAPNASTVIG 151 HALDT...

Further work revealed the complete nucleotide sequence <SEQ ID 261>:

1 ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT TCCGGCTGGT 51 GCTTTTTCATATCCTCATCA TCGCCGCCAG CAACTATCTG GTGCAGTTCC 101 CTTTCCAAAT TTTCGGCATCCACACCACTT GGGGCGCATT TTCCTTTCCC 151 TTCATCTTCC TTGCCACCGA CCTGACCGTCCGCATTTTCG GTTCTCACTT 201 GGCACGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTTTTGCTTTCCT 251 ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACAGG CTTGGGCGCG301 CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCCTTAG CCAGCTTTGC 351CGCCTACGCG ATCGGACAAA TCCTTGATAT TTTTGTATTC AACAAATTAC 401 GCCGTCTGAAAGCGTGGTGG ATTGCACCGA CCGCATCAAC CGTCATCGGC 451 AACGCCTTGG ATACGCTGGTATTTTTCGCC GTTGCCTTCT ACGCAAGCAG 501 CGATGGATTT ATGGCGGCAA ACTGGCAGGGCATCGCTTTT GTCGATTACC 551 TGTTCAAACT TACCGTCTGC ACCCTCTTCT TCCTGCCCGCCTACGGCGTG 601 ATACTGAATC TGCTGACGAA AAAACTGACA ACCCTGCAAA CCAAACAGGC651 GCAAGACCGC CCCGCGCCCT CGCTGCAAAA TCCGTAA

This corresponds to the amino acid sequence <SEQ ID 262; ORF66-1>:

1 MYAFTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFQIFGI HTTWGAFSFP 51FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF HNGSWTGLGA 101LSEFNTFVGR IALASFAAYA IGQILDIFVF NKLRRLKAWW IAPTASTVIG 151NALDTLVFFA VAFYASSDGF MAANWQGIAF VDYLFKLTVC TLFFLPAYGV 201ILNLLTKKLT TLQTKQAQDR PAPSLQNP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Hypothetical Protein o221 of E. coli (Accession NumberP37619)

ORF66 and o221 protein show 67% aa identity in 155aa overlap:

orf66 1 MYAFTAAQQQKALFRLVLFHILIIAASNYLVQFPFQIFGIHTTWGAFSFPFIFLATDLTV 60M  F+  Q+ KALF L LFH+L+I +SNYLVQ P  I G HTTWGAFSFPFIFLATDLTV o221 1MNVFSQTQRYKALFWLSLFHLLVITSSNYLVQLPVSILGFHTTWGAFSFPFIFLATDLTV 60 orf66 61RIFGSHLARRIIFWVMFPALLLSYVFSVLFHNGSWTGLGALSEFNTFVGRIALASFAAYA 120 RIFG+LARRIIF VM PALL+SYV S LF+ GSW G GAL+ FN FV RIA ASF AYA o221 61RIFGAPLARRIIFAVMIPALLISYVISSLFYMGSWQGFGALAHFNLFVARIATASFMAYA 120 orf66121 IGQILDIFVFNKLRRLKAWWIAPNASTVIGHALDT 155 +GQILD+ VFN+LR+ + WW+AP AST+G+  DT o221 121 LGQILDVHVFNRLRQSRRWWLAPTASTLFGNVSDT 155Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF66 shows 96.1% identity over a 155aa overlap with an ORF (ORF66a)from strain A of N. meningitidis:

The complete length ORF66a nucleotide sequence <SEQ ID 263> is:

1 ATGTACGCAT TTACCGCCGC ACAGCAACAG AAGGCACTCT TCTGGCTGGT 51GCTTTTTCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG GTGCAGTTCC 101CCTTCCAAAT TTCCGGCATC CACACCACTT GGGGCGCGTT TTCCTTTCCC 151TTCATCTTCC TCGCCACCGA CCTGACCGTC CGCATTTTCG GTTCGCACTT 201GGCACGGCGG ATTATCTTTT GGGTCATGTT CCCCGCCCTT TTGCTTTCCT 251ACGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACGGG CTTGGGCGCG 301CTGTCCGAAT TCAACACCTT TGTCGGACGC ATCGCGCTGG CAAGTTTTGC 351CGCCTACGCG CTCGGACAAA TCCTTGATAT TTTTGTGTTC AACAAATTAC 401GCCGTCTGAA AGCGTGGTGG GTTGCCCCGA CTGCATCAAC CGTCATCGGC 451AACGCCTTAG ATACGTTGGT ATTTTTCGCC GTTGCCTTCT ACGCAAGCAG 501CGATGGATTT ATGGCGGCAA ACTGGCAGGG CATCGCTTTT GTCGATTACC 551TGTTCAAACT CACCGTCTGC GGTCTGTTTT TCCTGCCCGC CTACGGCGTG 601ATTCTGAATC TGCTGACGAA AAAACTGACG ACCCTGCAAA CCAAACAGGC 651GCAAGACCGC CCCGCGCCCT CGCTGCAAAA TCCGTAA

This encodes a protein having amino acid sequence <SEQ ID 264>:

1 MYAFTAAQQQ KALFWLVLFH ILIIAASNYL VQFPFQISGI HTTWGAFSFP 51FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF HNGSWTGLGA 101LSEFNTFVGR IALASFAAYA LGQILDIFVF NKLRRLKAWW VAPTASTVIG 151NALDTLVFFA VAFYASSDGF MAANWQGIAF VDYLFKLTVC GLFFLPAYGV 201ILNLLTKKLT TLQTKQAQDR PAPSLQNP*

ORF66a and ORF66-1 show 97.8% identity in 228 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF66 shows 94.2% identity over a 155aa overlap with a predicted ORF(ORF66.ng) from N. gonorrhoeae:

The complete length ORF66ng nucleotide sequence <SEQ ID 265> is:

1 ATGTACGCAT TGACCGCCGC ACAGCAACAG AAGGCACTCT TCCGGCTGGT 51GCTTTTCCAT ATCCTCATCA TCGCCGCCAG CAACTATCTG GTGCAGTTCC 101CCTTCCGGAT TTTCGGCATC CACACCACTT GGGGCGCGTT TTCCTTTCCC 151TTCATCTTCC TCGCCACCGA CCTGACCGTC CGCATTTTCG GTTCGCACTT 201GGCGCGGCGG ATTATCTTTT GGGTGATGTT CCCCGCCCTT ttgCTTTcat 251aCGTCTTTTC CGTTTTGTTC CACAACGGCA GTTGGACGGG CTTGGGCGCG 301ctgTCCCAAT TCAACACCTT TGTCGGACGC ATCGCGCTGG CAAGTTTTGC 351CGCCTACGCG CTCGGACAAA TCCTTGATAT TTTCGTATTC GACAAATTAC 401GCCGTCTGAA AGCGTGGTGG ATTGCCCCGG CCGCATCAAC CGTCATCGGC 451AATGCACTGG ACACGTTAGT ATTTTTTGCC GTTGCCTTTT ACGCAAGCAG 501CGATGAATTT ATGGCGGCAA ACTGGCAGGG CATCGCTTTT GTCGATTACC 551TGTTCAAACT TACCGTCTGC ACCCTCTTCT TCCTGCCCGC CTACGGCGTG 601ATACTGAATC TGCTGACGAA AAAACTGACG GCCCTGCAAA CCAAACAGGC 651GCAAGACCGC CCCGTGCCCT CGCTGCAAAA TCCGTAA

This encodes a protein having amino acid sequence <SEQ ID 266>:

1 MYALTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFRIFGI HTTWGAFSFP 51FIFLATDLTV RIFGSHLARR IIFWVMFPAL SLSYVFSVLF HNGSWTGLGA 101PSQFNTFVGR IALASFAAYA LGQILDIFVF DKLRRLKAWW IAPAASTVIG 151NALDTLVFFA VAFYASSDEF MAANWQGIAF VDYLFKLTVC TLFFLPAYGV 201ILNLLTKKLT ALQTKQAQDR PVPSLQNP*

An alternative annotated sequence is:

1 MYALTAAQQQ KALFRLVLFH ILIIAASNYL VQFPFRIFGI HTTWGAFSFP 51FIFLATDLTV RIFGSHLARR IIFWVMFPAL LLSYVFSVLF HNGSWTGLGA 101LSQFNTFVGR IALASFAAYA LGQILDIFVF DKLRRLKAWW IAPAASTVIG 151NALDTLVFFA VAFYASSDEF MAANWQGIAF VDYLFKLTVC TLFFLPAYGV 201ILNLLTKKLT ALQTKQAQDR PVPSLQNP*

ORF66ng and ORF66-1 show 96.1% identity in 228 aa overlap:

Furthermore, ORF66ng shows significant homology with an E. coli ORF:

sp|P37619|YHHQ_ECOLI HYPOTHETICAL 25.3 KD PROTEIN IN FTSY-NIKA INTERGENICREGION (O221)>gi|1073495|pir||S47690 hypothetical protein o221 - Escherichia coli>gi|466607 (U00039) No definition line found [Escherichia coli] >gi|1789882 (AE000423) hypothetical 25.3 kD protein in ftsY-nikAintergenic region [Escherichia coli] Length = 221 Score =273 bits (692), Expect = 5e−73 Identities = 132/203 (65%), Positives =155/203 (76%) Query: 1MYALTAAQQQKALFRLVLFHILIIAASNYLVQFPFRIFGIHTTWGAFSFPFIFLATDLTV 60 M   + Q+ KALF L LFH+L+I +SNYLVQ P  I G HTTWGAFSFPFIFLATDLTV Sbjct: 1MNVFSQTQRYKALFWLSLFHLLVITSSNYLVQLPVSILGFHTTWGAFSFPFIFLATDLTV 60 Query:61 RIFGSHLARRIIFWVMFPALLLSYVFSVLFHNGSWTGLGALSQFNTFVGRIALASFAAYA 120RIFG+ LARRIIF VM PALL+SYV S LF+ GSW G GAL+ FN FV RIA ASF AYA Sbjct: 61RIFGAPLARRIIFAVMIPALLISYVISSLFYMGSWQGFGALAHFNLFVARIATASFMAYA 120 Query:121 LGQILDIFVFDKLRRLKAWWIAPAASTVIGNALDTLVFFAVAFYASSDEFMAANWQGIAF 180LGQILD+ VF++LR+ + WW+AP AST+ GN  DTL FF +AF+ S D FMA +W  IA Sbjct: 121LGQILDVHVFNRLRQSRRWWLAPTASTLFGNVSDTLAFFFIAFWRSPDAFMAEHWMEIAL 180 Query:181 VDYLFKLTVCTLFFLPAYGVILN 203 VDY FK+ +  +FFLP YGV+LN Sbjct: 181VDYCFKVLISIVFFLPMYGVLLN 203

Based on this analysis, including the homology with the E. coli proteinand the presence of several putative transmembrane domains in thegonococcal protein, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 32

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 267>:

1 ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT CGATAATTGC 51AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAAyGCA GTmwrAATAT 101CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT TCATAAGTTT 151GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA AAACGGTAGA 201TTTAACACAC AyyCCTACGG GCGCAAAAGC CCGAATCAAC GCCAAAATAA 251CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG CAAACTTGCC 301CGCTTAGgCG CGAAATTCAG CACAAGGGCG GTtCCCTATG TCGGAACAGC 351CcTTTTAGCC CACGACGTAT ACGAAAcTTT CAAAGAAGAC ATACAGGCAC 401GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGTAAA AGGCTACGAA 451TATAGTAATT GCCTTTGGTA CGAAGACAAA AGACGTATTA ATAGAACCTA 501TGGCTGCTAC GGCGTTGAT..

This corresponds to the amino acid sequence <SEQ ID 268; ORF72>:

1 MVIKYTNLNF AKLSIIAILM MYSFEANANA VXISETVSVD TGQGAKIHKF 51VPKNSKTYSS DLIKTVDLTH XPTGAKARIN AKITASVSRA GVLAGVGKLA 101RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP ETDKFVKGYE 151YSNCLWYEDK RRINRTYGCY GVD..

Further work revealed the complete nucleotide sequence <SEQ ID 269>:

1 ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT CGATAATTGC 51AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA GTAAAAATAT 101CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT TCATAAGTTT 151GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA AAACGGTAGA 201TTTAACACAC ATCCCTACGG GCGCAAAAGC CCGAATCAAC GCCAAAATAA 251CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG CAAACTTGCC 301CGCTTAGGCG CGAAATTCAG CACAAGGGCG GTTCCCTATG TCGGAACAGC 351CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC ATACAGGCAC 401GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGCAAA GGTCTCAGGC 451 TAA

This corresponds to the amino acid sequence <SEQ ID 270; ORF72-1>:

1 MVIKYTNLNF AKLSIIAILM MYSFEANANA VKISETVSVD TGQGAKIHKF 51VPKNSKTYSS DLIKTVDLTH IPTGAKARIN AKITASVSRA GVLAGVGKLA 101RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP ETDKFAKVSG 151 *

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF72 shows 98.0% identity over a 147aa overlap with an ORF (ORF72a)from strain A of N. meningitidis:

The complete length ORF72a nucleotide sequence <SEQ ID 271> is:

1 ATGGTCATAA AATATACAAA TTTGAATTTT GCGAAATTGT CGATAATTGC 51AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA GTAAAAATAT 101CTGAAACTGT TTCAGTTGAT ACCGGACAAG GTGCGAAAAT TCATAAGTTT 151GTACCTAAAA ATAGTAAAAC TTATTCATCT GATTTAATAA AAACGGTAGA 201TTTAACACAC ATCCCTACGG GCGCAAAAGC CCGAATCAAC GCCAAAATAA 251CCGCCAGCGT ATCCCGCGCC GGCGTATTGG CGGGGGTCGG CAAACTTGCC 301CGCTTAGGCG CGAAATTCAG CACAAGGGCG GTTCCCTATG TCGGAACAGC 351CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC ATACAGGCAC 401GAGGCTACCA ATACGACCCC GAAACCGACA AATTTGCAAA GGTCTCAGGC 451 TAA

This encodes a protein having amino acid sequence <SEQ ID 272>:

1 MVIKYTNLNF AKLSIIAILM MYSFEANANA VKISETVSVD TGQGAKIHKF 51VPKNSKTYSS DLIKTVDLTH IPTGAKARIN AKITASVSRA GVLAGVGKLA 101RLGAKFSTRA VPYVGTALLA HDVYETFKED IQARGYQYDP ETDKFAKVSG 151 *

ORF72a and ORF72-1 show 100.0% identity in 150 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF72 shows 89% identity over a 173aa overlap with a predicted ORF(ORF72.ng) from N. gonorrhoeae:

An ORF72ng nucleotide sequence <SEQ ID 273> was predicted to encode aprotein having amino acid sequence <SEQ ID 274>:

1 MVTKHTNLNF AKLSIIAILM MYSFEANANA VKISETLSVD TGQGAKVHKF 51VPKSSNIYSS DLTKAVDLTH IPTGAKARIN AKITASVSRA GVLSGVGKLV 101RQGAKFGTRA VPYVGTALLA HDVYETFKED IQARGCRYDP ETDKFVKGYE 151YANCLWYEDE RRINRTYGCY GVDSSIMRLM PDRSRFPEVK QLMESQMYRL 201ARPFWNWRKE ELNKLSSLDW NNFVLNRCTF DWNGGGCAVN KGDDFRAGAS 251FSLGRNPKYK EEMDAKKPEE ILSLKVDADP DKYIEATGYP GYSEKVEVAP 301GTKVNMGPVT DRNGNPVQVA ATFGRDAQGN TTADVQVIPR PDLTPASAEA 351PHAQPLPEVS PAENPANNPD PDENPGTRPN PEPDPDLNPD ANPDTDGQPG 401TSPDSPAVPD RPNGRHRKER KEGEDGGLSC DYFPEILACQ EMGKPSDRMF 451HDISIPQVTD DKTWSSHNFL PSNGVCPQPK TFHVFGRQYR ASYEPLCVFA 501EKIRFAVLLA FIIMSAFVVF GSLGGE*

After further analysis, the following gonococcal DNA sequence <SEQ ID275> was identified:

1 ATGGTCACAA AACATACAAA TTTGAATTTT GCGAAATTGT CGATAATTGC 51AATTTTGATG ATGTATTCGT TTGAAGCGAA TGCAAATGCA GTAAAAATAT 101CTGAAACTCT TTCGGTTGAT ACCGGACAAG GCGCGAAAGT TCATAAGTTC 151GTTCCTAAAT CAAGTAATAT TTATTCATCT GATTTAACAA AAGCGGTAGA 201TTTAACGCAT ATCCCCACGG GCGCAAAAGC CCGAATCAAC GCCAAAATAA 251CCGCCAGCGT ATCCCGCGCC GGCGTATTGT CGGGGGTCGG CAAACTTGTC 301CGCCAAGGCG CGAAATTCGG CACAAGGGCG GTTCCCTATG TCGGAACAGC 351CCTTTTAGCC CACGACGTAT ACGAAACTTT CAAAGAAGAC ATACAGGCAC 401GAGGCTGCCG ATACGATCCC GAAACCGACA AATTT

This corresponds to the amino acid sequence <SEQ ID 276; ORF72ng-1>:

1 MVTKHTNLNF AKLSIIAILM MYSFEANANA VKISETLSVD TGQGAKVHKF 51VPKSSNIYSS DLTKAVDLTH IPTGAKARIN AKITASVSRA GVLSGVGKLV 101RQGAKFGTRA VPYVGTALLA HDVYETFKED IQARGCRYDP ETDKF

ORF72ng-1 and ORF721-1 show 89.7% identity in 145 aa overlap:

Based on this analysis, including the presence of a putative leadersequence and transmembrane domains in the gonococcal protein, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 33

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 277>:

1 ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT TGGAGATTAT 51GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGCTGG ACGTTGTTTT 101TGATGGCGGC AGGTTTTGCC GCCGGCGTGC TGATGCTCAG GCAAACCGGG 151GCTGACCGGT CTTTTATTGG CGGGCGCGGC AATGAGAAGC GGCGGGAAGG 201TATCCGTTTA TCAGATGTTG TGGCCTATC..

This corresponds to the amino acid sequence <SEQ ID 278; ORF73>:

1 MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAAGFA AGVLMLRQTG 51LTGLLLAGAA MRSGGKVSVY QMLWPI..

Further work revealed the complete nucleotide sequence <SEQ ID 279>:

1 ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT TGGAGATTAT 51GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGCTGG ACGTTGTTTT 101TGATGGCGGC AGGTTTTGCC GCCGGCGTGC TGATGCTCAG GCATACGGGG 151CTGTCCGGTC TTTTATTGGC GGGCGCGGCA ATGAGAAGCG GCGGGAGGGT 201ATCCGTTTAT CAGATGTTGT GGCCTATCCG TTATACGGTG GCGGCTGTGT 251GTCTGATGAG TCCGGGATTC GTATCCTCGG TGTTGGCGGT ATTGCTGCTG 301CTGCCGTTTA AGGGAGGGGC AGTGTTGCAG GCAGGAGGTG CGGAAAATTT 351TTTCAACATG AACCAATCGG GCAGAAAAGA GGGCTTTTCC CGCGATGACG 401ATATTATCGA GGGAGAATAT ACGGTTGAAG AGCCTTACGG CGGCAATCGT 451TCCCGAAACG CCATCGAACA CAAAAAAGAC GAATAA

This corresponds to the amino acid sequence <SEQ ID 280; ORF73-1>:

1 MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAAGFA AGVLMLRHTG 51LSGLLLAGAA MRSGGRVSVY QMLWPIRYTV AAVCLMSPGF VSSVLAVLLL 101LPFKGGAVLQ AGGAENFFNM NQSGRKEGFS RDDDIIEGEY TVEEPYGGNR 151 SRNAIEHKKD E*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF73 shows 90.8% identity over a 76aa overlap with an ORF (ORF73a) fromstrain A of N. meningitidis:

The complete length ORF73a nucleotide sequence <SEQ ID 281> is:

1 ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT TGGAGATTAT 51GTCGATTGTG TGGGTTGCCG ATTGGTTGGG CGGCGGTTGG ACGCTGTTTC 101TAATGGCGGC AACCTTTGCC GCCGGCGTGG TGATGCTCAG GCATACGGGG 151CTGTCCGGTC TTTTATTGGC GGGCGCGGCA ATGAGAAGCG GCGGGAGGGT 201ATCCGTTTAT CANATGTTGT GGCNTATCCG TTATACGGTG GCGGCGGTGT 251GTCNGATGAG TCCGGGATTC GTATCCTCGG TGTNGGCGGT ATTGCTGNTG 301CTNCCGTTTA AGGGAGGTGC AGTGTTGCAG GCAGGAGGTG CGGAAAATTT 351TTTCAACATG AACCANTCGG GCAGAAAAGA NGGCNTTTCC CGCGATGACG 401ATATTATCGA GGGGGAATAT ACGGTTGAAG ANCCTTACGG CGGCANTCGT 451TTCCGAAACG CCNTNGAACA CAAAAAAGAC GAATAA

This encodes a protein having amino acid sequence <SEQ ID 282>:

1 MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAATFA AGVVMLRHTG 51LSGLLLAGAA MRSGGRVSVY XMLWXIRYTV AAVCXMSPGF VSSVXAVLLX 101LPFKGGAVLQ AGGAENFFNM NXSGRKXGXS RDDDIIEGEY TVEXPYGGXR 151 FRNAXEHKKD E*

ORF73a and ORF73-1 show 91.3% identity in 161 aa overlap

Homology with a Predicted ORF from N. gonorrhoeae

ORF73 shows 92.1% identity over a 76aa overlap with a predicted ORF(ORF73.ng) from N. gonorrhoeae:

The complete length ORF73ng nucleotide sequence <SEQ ID 283> is:

1 ATGAGATTTT TCGGTATCGG TTTTTTGGTG CTGCTGTTTT TGGAAATTAT 51GTCGATTGTG TGGGTTGCCG ATTGGCTGGG CGGCGGTTGG AcgcTGTTTC 101TAATGGCGGC AACCTTTGCC GCCGGTGTGC TGATGCTCAG GCATAcggGG 151CTGTCCGGTC TTTTATTGGC TGGCGCGGCG GTAAAAagta gtgGGAAGGT 201ATCTGTTTAT CagatgtTGT GGCCTATCCG TTATAcggtg gcggcggtgT 251GTCTGatgag tCcggGATTC GTATCCTccg tgttggCGGT ATTGCTGCTG 301CTGCcgttta aggGaggGgc agtgttgcag gcaggaggtg cggaaaATTT 351TTTCAACATg aaCcaatcgg gcagaaAaga gggatttttc cacgatgacg 401atattatcga gggagaatat acggttgaaa aacctgacgg cggcaatcgt 451tcccgaAAcg ccatcgaaca cgaaaAagac gaataA

This encodes a protein having amino acid sequence <SEQ ID 284>:

1 MRFFGIGFLV LLFLEIMSIV WVADWLGGGW TLFLMAATFA AGVLMLRHTG 51LSGLLLAGAA VKSSGKVSVY QMLWPIRYTV AAVCLMSPGF VSSVLAVLLL 101LPFKGGAVLQ AGGAENFFNM NQSGRKEGFF HDDDIIEGEY TVEKPDGGNR 151 SRNAIEHEKD E*

ORF73ng and ORG73-1 show 93.8% identity in 161 aa overlap

Based on this analysis, including the presence of a putative leadersequence and putative transmembrane domain in the gonococcal protein, itis predicted that the proteins from N. meningitidis and N. gonorrhoeae,and their epitopes, could be useful antigens for vaccines ordiagnostics, or for raising antibodies.

Example 34

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 285>:

1 ATGTTTGTTT TTCAGACGGC ATTCTT.ATG TTTCAGAAAC ATTTGCAGAA 51AGCCTCCGAC AGCGTCGTCG GAGGGACATT ATACGTGGTT GCCACGCCCA 101TCGGCAATTT GGCGGACATT ACCCTGCGCG CTTTGGCGGT ATTGCAAAAG 151GCG....... .....GCCGA AGACACGCGC GTTACCGCAC AGCTTTTGAG 201CGCGTACGGC ATTCAGGGCA AACTCGTCAG TGTGCGCGAA CACAACGAAC 251GGCAGATGGC GGACAAGATT GTCGGCTATC TTTCAGACGG CATGGTTGTG 301GCACAGGTTT CCGATGCGGG TACGCCGGCC GTGTGCGACC CGGGCGCGAA 351ACTCGCCCGC CGCGTGCGTG AGGCCGGGTT TAAAGTCGTT CCCGTCGTGG 401GCGCAAC.GC GGTGATGGCG GCTTTGAGCG TGGCCGGTGT GGAAGGATCC 451GATTTTTATT TCAACGGTTT TGTACCGCCG AAATCGGGAG AACGCAGGAA 501ACTGTTTGCC AAATGGGTGC GGGCGGCGTT TCCTATCGTC ATGTTTGAAA 551CGCCGCACCG CATCGGTGCA GCGCTTGCCG ATATGGCGGA ACTGTTCCCC 601GAACGCCGAT TAATGCTGGC GCGCGAAATT ACGAAAACGT TTGAAACGTT 651CTTAAGCGGC ACGGTTGGGG AAATTCAGAC GGCATTGTCT GCCGACGGCG 701ACCAATCGCG CGGCGAGATG GTGTTGGTGC TTTATCCGGC GCAGGATGAA 751AAACACGAAG GCTTGTCCGA GTCCGCGCAA AACATCATGA AAATCCTCAC 801AGCCGAGCTG CCGACCAAAC AGGCGGCGGA GCTTGCTGCC AAAATCACGG 851GCGAGGGAAA GAAAGCTTTG TACGAT..

This corresponds to the amino acid sequence <SEQ ID 286; ORF75>:

1 MFVFQTAFXM FQKHLQKASD SVVGGTLYVV ATPIGNLADI TLRALAVLQK 51A....AEDTR VTAQLLSAYG IQGKLVSVRE HNERQMADKI VGYLSDGMVV 101AQVSDAGTPA VCDPGAKLAR RVREAGFKVV PVVGAXAVMA ALSVAGVEGS 151DFYFNGFVPP KSGERRKLFA KWVRAAFPIV MFETPHRIGA ALADMAELFP 201ERRLMLAREI TKTFETFLSG TVGEIQTALS ADGDQSRGEM VLVLYPAQDE 251KHEGLSESAQ NIMKILTAEL PTKQAAELAA KITGEGKKAL YD..

Further work revealed the complete nucleotide sequence <SEQ ID 287>:

1 ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG TCGGAGGGAC 51ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCGGAC ATTACCCTGC 101GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATCTGTGC CGAAGACACG 151CGCGTTACCG CACAGCTTTT GAGCGCGTAC GGCATTCAGG GCAAACTCGT 201CAGTGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAG ATTGTCGGCT 251ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC GGGTACGCCG 301GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC GTGAGGCCGG 351GTTTAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTGATG GCGGCTTTGA 401GCGTGGCCGG TGTGGAAGGA TCCGATTTTT ATTTCAACGG TTTTGTACCG 451CCGAAATCGG GAGAACGCAG GAAACTGTTT GCCAAATGGG TGCGGGCGGC 501GTTTCCTATC GTCATGTTTG AAACGCCGCA CCGCATCGGT GCGACGCTTG 551CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCT GGCGCGCGAA 601ATTACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG GGGAAATTCA 651GACGGCATTG TCTGCCGACG GCAACCAATC GCGCGGCGAG ATGGTGTTGG 701TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC CGAGTCCGCG 751CAAAACATCA TGAAAATCCT CACAGCCGAG CTGCCGACCA AACAGGCGGC 801GGAGCTTGCT GCCAAAATCA CGGGCGAGGG AAAGAAAGCT TTGTACGATC 851TGGCTCTGTC TTGGAAAAAC AAATAG

This corresponds to the amino acid sequence <SEQ ID 288; ORF75-1>:

1 MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ KADIICAEDT 51RVTAQLLSAY GIQGKLVSVR EHNERQMADK IVGYLSDGMV VAQVSDAGTP 101AVCDPGAKLA RRVREAGFKV VPVVGASAVM AALSVAGVEG SDFYFNGFVP 151PKSGERRKLF AKWVRAAFPI VMFETPHRIG ATLADMAELF PERRLMLARE 201ITKTFETFLS GTVGEIQTAL SADGNQSRGE MVLVLYPAQD EKHEGLSESA 251QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN K*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF75 shows 95.8% identity over a 283aa overlap with an ORF (ORF75a)from strain A of N. meningitidis:

The complete length ORF75a nucleotide sequence <SEQ ID 289> is:

1 ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG TCGGAGGGAC 51ATTATACGTG GTTGCCACGC CCATCGGCAA TTTGGCGGAC ATTACCCTGC 101GCGCTTTGGC GGTATTGCAA AAGGCGGACA TCATCTGTGC CGAAGACACG 151CGCGTTACCG CGCAGCTTTT GAGCGCGTAC GGCATTCAGG GCAAACTCGT 201CAGCGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAG ATTGTCGGCT 251ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC GGGTACGCCG 301GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC GTGAGGTCGG 351GTTTAAAGTT GTCCCTGTTG TCGGCGCAAG CGCGGTGATG GCGGCTTTGA 401GTGTGGCTGG TGTGGCGGGA TCCGATTTTT ATTTCAACGG TTTTGTACCG 451CCGAAATCGG GCGAACGTAG GAAATTGTTT GCCAAATGGG TGCGGGTGGC 501GTTTCCCGTC GTGATGTTTG AAACGCCGCA CCGCATCGGG GCGACGCTTG 551CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCT GGCGCGCGAA 601ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG GGGAAATTCA 651GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG ATGGTGTTGG 701TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC CGAGTCCGCG 751CAAAACATCA TGAAAATCCT CACAGCCGAG CTGCCGACCA AACAGGCGGC 801GGAGCTTGCC GCCAAAATCA CGGGCGAGGG AAAAAAAGCT TTGTACGATC 851TGGCACTGTC TTGGAAAAAC AAATGA

This encodes a protein having amino acid sequence <SEQ ID 290>:

1 MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ KADIICAEDT 51 RVTAQLLSAYGIQGKLVSVR EHNERQMADK IVGYLSDGMV VAQVSDAGTP 101 AVCDPGAKLARRVREVGFKV VPVVGASAVM AALSVAGVAG SDFYFNGFVP 151 PKSGERRKLF AKWVRVAFPVVMFETPHRIG ATLADMAELF PERRLMLARE 201 ITKTFETFLS GTVGEIQTAL AADGNQSRGEMVLVLYPAQD EKHEGLSESA 251 QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN K*

ORF75a and ORF75-1 show 98.3% identity in 291 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF75 shows 93.2% identity over a 292aa overlap with a predicted ORF(ORF75.ng) from N. gonorrhoeae:

An ORF75ng nucleotide sequence <SEQ ID 291> was predicted to encode aprotein having amino acid sequence <SEQ ID 292>:

1 MSVFQTAFFM FQKHLQKASD SVVGGTLYVV ATPIGNLADI TLRALAVLQK 51 ADIICAEDTRVTAQLLSAYG IQGRLVSVRE HNERQMADKV IGFLSDGLVV 101 AQVSDAGTPA VCDPGAKLARRVREAGFKVV PVVGASAVMA ALSVAGVAES 151 DFYFNGFVPP KSGERRKLFA KWVRAAFPVVMFETPHRIGA TLADMAELFP 201 ERRLMLAREI TKTFETFLSG TVGEIQTALA ADGNQSRGEMVLVLYPAQDE 251 KHEGLSESAQ NAMKILAAEL PTKQAAELAA KITGEGKKAL YDLALSWKNK301 *

After further analysis, the following gonococcal DNA sequence <SEQ ID293> was identified:

1 ATGTTTCAGA AACACTTGCA GAAAGCCTCC GACAGCGTCG TCGGAGGGAC 51 ATTATACGTGGTTGCCACGC CCATCGGCAA TTTGGCAGAC ATTACCCTGC 101 GCGCTTTGGC GGTATTGCAAAAGGCGGACA TCATTTGTGC CGAAGACACG 151 CGCGTTACTG CGCAGCTTTT GAGCGCGTACGGCATTCAGG GCAGGTTGGT 201 CAGTGTGCGC GAACACAACG AGCGGCAGAT GGCGGACAAGGTAATCGGTT 251 TCCTTTCAGA CGGCCTGGTT GTGGCGCAGG TTTCCGATGC GGGTACGCCG301 GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC GCGAAGCAGG 351GTTCAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTAATG GCGGCGTTGA 401 GTGTGGCCGGTGTGGCGGAA TCCGATTTTT ATTTCAACGG TTTTGTACCG 451 CCGAAATCGG GCGAACGTAGGAAATTGTTT GCCAAATGGG TGCGGGCGGC 501 ATTTCCTGTC GTCATGTTTG AAACGCCGCACCGAATCGGG GCAACGCTTG 551 CCGATATGGC GGAATTGTTC CCCGAACGCC GTCTGATGCTGGCGCGCGAA 601 ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG GGGAAATTCA651 GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG ATGGTGTTGG 701TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC CGAGTCTGCG 751 CAAAATGCGATGAAAATCCT TGCGGCCGAG CTGCCGACCA AGCAGGCGGC 801 GGAGCTTGCC GCCAAGATTACAGGTGAGGG CAAAAAGGCT TTGTACGATT 851 TGGCACTGTC GTGGAAAAAC AAATGA

This corresponds to the amino acid sequence <SEQ ID 294; ORF75ng-1>:

1 MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ KADIICAEDT 51 RVTAQLLSAYGIQGRLVSVR EHNERQMADK VIGFLSDGLV VAQVSDAGTP 101 AVCDPGAKLARRVREAGFKV VPVVGASAVM AALSVAGVAE SDFYFNGFVP 151 PKSGERRKLF AKWVRAAFPVVMFETPHRIG ATLADMAELF PERRLMLARE 201 ITKTFETFLS GTVGEIQTAL AADGNQSRGEMVLVLYPAQD EKHEGLSESA 251 QNAMKILAAE LPTKQAAELA AKITGEGKKA LYDLALSWKN K*

ORF75ng-1 and ORF75-1 show 96.2% identity in 291 aa overlap:

Furthermore, ORG75ng-1 shows significant homology to a hypothetical E.coli protein:

sp|P45528|YRAL_ECOLI HYPOTHETICAL 31.3 KD PROTEIN IN AGAI-MTR INTERGENICREGION (F286) >gi|606086 (U18997) ORF_f286 [Escherichiacoli] >gi|1789535 (AE000395) hypothetical 31.3 kD protein in agai-mtrintergenic region [Escherichia coli] Length = 286 Score = 218 bits(550), Expect = 3e−56 Identities = 128/284 (45%), Positives = 171/284(60%), Gaps = 4/284 (1%) Query: 4KHLQKASDSVVGGTLYVVATPIGNLADITLRALAVLQKADIICAEDTRVTAQLLSAYGIQ 63 K  Q A+S   G LY+V TPIGNLADIT RAL VLQ  D+I AEDTR T  LL  +GI Sbjct: 2KQHQSADNSQ--GQLYIVPTPIGNLADITQRALEVLQAVDLIAAEDTRHTGLLLQHFGIN 59 Query:64 GRLVSVREHNERQMADKVIGFLSDGLVVAQVSDAGTPAVCDPGAKLARRVREAGFKVVPV 123  RL++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG  L R  REAG +VVP+ Sbjct: 60ARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPGYHLVRTCREAGIRVVPL 119 Query:124 VGASAVMAALSVAGVAESDFYFNGFVPPKSGERRKLFAKWVRAAFPVVMFETPHRIGATL 183 G  A + ALS AG+    F + GF+P KS  RR            ++ +E+ HR+  +L Sbjct: 120PGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAEPRTLIFYESTHRLLDSL 179 Query:184 ADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALAADGNQSRGEMVLVLYPAQDEK 242 D+  +  E R ++LARE+TKT+ET     VGE+   +  D N+ +GEMVL++      + Sbjct: 180EDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDENRRKGEMVLIV-EGHKAQ 238 Query:243 HEGLSESAQNAMKILAAELPTKQAAELAAKITGEGKKALYDLAL 286  E L   A   + +LAELP K+AA LAA+I G  K ALY  AL Sbjct: 239EEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALYKYAL 282

Based on this analysis, including the presence of a putativetransmembrane domain in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 35

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 295>:

This corresponds to the amino acid sequence <SEQ ID 296; ORF76>:

Further work revealed the complete nucleotide sequence <SEQ ID 297>:

1 ATGAAACAGA AAAAAACCGC TGCCGCAGTT ATTGCTGCAA TGTTGGCAGG 51 TTTTGCGGCAGCCAAAGCAC CCGAAATCGA CCCGGCTTTG GTGGATACGC 101 TGGTGGCGCA GATCATGCAGCAGGCAGACC GGCATGCGGA GCAGTCCCAA 151 AAACCGGACG GGCAGGCAAT CCGAAACGATGCCGTCCGCC GGCTACAAAC 201 TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTGGATAAGGATA 251 AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT TTATGCCGAG301 GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG AAGACGAGCT 351GCACAAGTTT TACGAACAGC AAATCCGCAT GATCAAATTG CAGCAGGTCA 401 GCTTCGCAACCGAAGAGGAG GCGCGTCAGG CGCAGCAGCT CCTGCTCAAA 451 GGGCTGTCTT TTGAAGGGCTGATGAAGCGT TATCCGAACG ACGAGCAGGC 501 TTTTGACGGT TTCATTATGG CGCAGCAGCTTCCCGAGCCG CTGGCTTCGC 551 AGTTTGCCGC GATGAATCGG GGCGACGTTA CCCGCGATCCGGTCAAATTG 601 GGCGAACGCT ATTATCTGTT CAAACTCAGC GAGGTCGGGA AAAACCCCGA651 CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAGCAG GGTTTGAGAC 701AGGAAAAAGC CCGCTTGAAA ATCGATGCCC TTTTGGAAGA AAACGGTGTC 751 AAACCGTAA

This corresponds to the amino acid sequence <SEQ ID 298; ORF76-1>:

1 MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ QADRHAEQSQ 51 KPDGQAIRNDAVRRLQTLEV LKNRALKEGL DKDKDVQNRF KIAEASFYAE 101 EYVRFLERSE TVSEDELHKFYEQQIRMIKL QQVSFATEEE ARQAQQLLLK 151 GLSFEGLMKR YPNDEQAFDG FIMAQQLPEPLASQFAAMNR GDVTRDPVKL 201 GERYYLFKLS EVGKNPDAQP FELVRNQLEQ GLRQEKARLKIDALLEENGV 251 KP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF76 shows 96.7% identity over a 30aa overlap and 96.8% identity over a31aa overlap with an ORF (ORF76a) from strain A of N. meningitidis:

The complete length ORF76a nucleotide sequence <SEQ ID 299> is:

1 ATGAAACAGA AAAAAACCGC TGCCGCAGTT ATTGCTGCAA TGTTGGCAGG 51 TTTTGCGGCAGCCAAAGCAC CCGAAATCGA CCCGGCTTTG GTGGATACGC 101 TGGTGGCGCA GATCATGCAGCAGGCAGACC GGCATGCGGA GCAGTCCCAA 151 AAACCGGACG GGCAGGCAAT CCGAAACGATGCCGTCCGTC GGCTGCAAAC 201 TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTGGATAAGGATA 251 AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT TTATGCCGAG301 GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG AAAGCGCACT 351GCGTCAGTTT TATGAGCGGC AAATCCGCAT GATCAAATTG CAGCAGGTCA 401 GCTTCGCAACCGAAGAGGAG GCGCGTCAGG CGCAGCAGCT CCTGCTCAAA 451 GGGCTGTCTT TTGAAGGGCTGATGAAGCGT TATCCGAACG ACGAGCAGGC 501 TTTTGACGGT TTCATTATGG CGCAGCAGCTTCCCGAGCCG CTGGCTTCGC 551 AGTTTGCAGC GATGAATCGG GGCGACGTTA CCCGCGATCCGGTCAAATTG 601 GGCGAACGCT ATTATCTGTT CAAACTCAGC GAGGTCGGGA AAAACCCCGA651 CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAACAA GGTTTGAGAC 701AGGAAAAAGC CCGCTTGAAA ATCGATGCCA TTTTGGAAGA AAACGGTGTC 751 AAACCGTAA

This encodes a protein having amino acid sequence <SEQ ID 300>:

1 MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ QADRHAEQSQ 51 KPDGQAIRNDAVRRLQTLEV LKNRALKEGL DKDKDVQNRF KIAEASFYAE 101 EYVRFLERSE TVSESALRQFYERQIRMIKL QQVSFATEEE ARQAQQLLLK 151 GLSFEGLMKR YPNDEQAFDG FIMAQQLPEPLASQFAAMNR GDVTRDPVKL 201 GERYYLFKLS EVGKNPDAQP FELVRNQLEQ GLRQEKARLKIDAILEENGV 251 KP*

ORF76a and ORF76-1 show 97.6% identity in 252 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

The aligned aa sequences of ORF76 and a predicted ORF (ORF76.ng) from N.gonorrhoeae of the N- and C-termini show 96.7% and 100% identity in 30and 31 overlap, respectively:

The complete length ORF76ng nucleotide sequence <SEQ ID 301> is:

1 ATGAAACAGA AAAAGACCGC TGCCGCAGTT ATTGCTGCAA TGTTGGCAGG 51 TTTTGCGGCAGCCAAAGCAC CCGAAATCGA CCCGGCTTTG GTGGATACGC 101 TGGTGGCGCA GATCATGCAGCAGGCAGACC GGCATGCGGA GCAGTCCCAA 151 AGACCGGACG GGCAGGCAAT CCGAAACGATGCCGTCCGCC GGCTGCAAAC 201 TTTGGAAGTT TTGAAAAACA GGGCATTGAA GGAAGGTTTGGATAAGGATA 251 AGGATGTCCA AAACCGCTTT AAAATCGCCG AAGCGTCTTT TTATGCCGAG301 GAGTACGTCC GTTTTCTGGA ACGTTCGGAA ACGGTTTCCG AAAGCGCACT 351GCGTCAGTTT TATGAGCGGC AAATCCGCAT GATCAAATTG CAGCAGGTCA 401 GCTTCGCAACCGAAGAGGAG GCGCGTCAGG CGCAGCAGCT CCTGCTCAAA 451 GGGCTGTCTT TTGAAGGGCTGATGAAGCGT TATCCGAACG ACGAGCAGGC 501 GTTCGACGGT TTCATTATGG CGCAGCAGCTTCCCGAGCCG CTGGCTTcgc 551 agtttgCCGG TATGAACCGT GGCGACGTTA CCCGCAATCCGGTCAAATTG 601 GGCGAACGCT ATTACCTGTT CAAACTCGGC GCGGTCGGGA AAAACCCCGA651 CGCGCAGCCT TTCGAGTTGG TCAGAAACCA GTTGGAACAA GGTTTGAGGC 701AGGAAAAAGC CCGCTTGAAA ATCGATGCCC TTTTGGAaga Aaacggtgtc 751 AaacCGTAA

This encodes a protein having amino acid sequence <SEQ ID 302>:

1 MKQKKTAAAV IAAMLAGFAA AKAPEIDPAL VDTLVAQIMQ QADRHAEQSQ 51 RPDGQAIRNDAVRRLQTLEV LKNRALKEGL DKDKDVQNRF KIAEASFYAE 101 EYVRFLERSE TVSESALRQFYERQIRMIKL QQVSFATEEE ARQAQQLLLK 151 GLSFEGLMKR YPNDEQAFDG FIMAQQLPEPLASQFAGMNR GDVTRNPVKL 201 GERYYLFKLG AVGKNPDAQP FELVRNQLEQ GLRQEKARLKIDALLEENGV 251 KP*

ORF76ng and ORF76-1 show 96.0% identity in 252 aa overlap

Furthermore, ORF76ng shows significant homology to a B. subtilis exportprotein precursor:

sp|P24327|PRSA_BACSU PROTEIN EXPORT PROTEIN PRSAPRECURSOR >gi|98227|pir||S15269 33K lipoprotein - Bacillussubtilis >gi|39782 (X57271) 33 kDa lipoprotein [Bacillussubtilis] >gi|2226124|gnl|PID|e325181 (Y14077) 33 kDa lipoprotein[Bacillus subtilis] >gi|2633331|gnl|PID|e1182997 (Z99109) molecularchaperonin [Bacillus subtilis] Length = 292 Score = 50.4 bits (118),Expect = 1e−05 Identities = 48/199 (24%), Positives = 82/199 (41%), Gaps= 32/199 (16%) Query: 70VLKNRALKEGLDK-----DKDVQNRFKIAEASF----------YAEEYVRFLERSETVSE 114VL     ++ LDK     DK++ N+ K  +             Y ++Y++   + E +++ Sbjct: 53VLTQLVQEKVLDKKYKVSDKEIDNKLKEYKTQLGDQYTALEKQYGKDYLKEQVKYELLTQ 112 Query:115 SA-----------LRQFYERQIRMIKLQQVSFATEEEARQAQQLLLKGLSFEGLMKRYPN 163 A           +++++E     I+   +  A ++ A + ++ L KG  FE L K Y Sbjct: 113KAAKDNIKVTDADIKEYWEGLKGKIRASHILVADKKTAEEVEKKLKKGEKFEDLAKEYST 172 Query:164 DEQAFDG-----FIMAQQLPEPLASQFAAMNRGDVTRDPVKLGERYYLFKLSEVGKNPDA 218D  A  G     F    Q+ E  +     +  G+V+ DPVK    Y++ K +E     D Sbjct: 173DSSASKGGDLGWFAKEGQMDETFSKAAFKLKTGEVS-DPVKTQYGYHIIKKTEERGKYDD 231 Query:219 QPFELVRNQLEQGLRQEKA 237    EL    LEQ L    A Sbjct: 232MKKELKSEVLEQKLNDNAA 250

Based on this analysis, including the presence of a putative leadersequence and a RGD motif in the gonococcal protein, it was predictedthat the proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

ORF76-1 (27.8 kDa) was cloned in the pET vector and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 10A shows the results ofaffinity purification of the His-fusion protein, Purified His-fusionprotein was used to immunise mice, whose sera were used for Western blot(FIG. 10B), ELISA (positive result), and FACS analysis (FIG. 10C). Theseexperiments confirm that ORF76-1 is a surface-exposed protein, and thatit is a useful immunogen.

Example 36

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 303>:

This corresponds to the amino acid sequence <SEQ ID 304; ORF81>:

Further work revealed the complete nucleotide sequence <SEQ ID 305>:

1 ATGAAAAAAT CTTTCCTTAC GCTTGTTCTG TATTCGTCTT TACTTACCGC 51 CAGCGAAATTGCCTATCGCT TTGTATTTGG GATTGAAACC TTACCGGCGG 101 CAAAAATTGC GGAAACGTTTGCGCTGACAT TTGTGATTGC TGCGCTGTAT 151 CTGTTTGCGC GTTATAAGGT GACGCGTTTGTTGATTGCGG TGTTTTTTGC 201 GTTCAGCATT ATTGCCAACA ATGTGCATTA CGCGGTTTATCAAAGCTGGA 251 TGACGGGCAT CAATTATTGG CTGATGCTGA AAGAGGTTAC CGAAGTCGGC301 AGCGCGGGTG CGTCGATGTT GGATAAGTTG TGGCTGCCTG TGTTGTGGGG 351CGTGTTGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC CGCCGTAAGA 401 CGCATTTTTCTGCCGATATA CTGTTTGCCT TCCTAATGCT GATGATTTTC 451 GTGCGTTCGT TCGACACGAAACAAGAGCAC GGTATTTCGC CCAAACCGAC 501 ATACAGCCGC ATCAAAGCCA ATTATTTCAGCTTCGGTTAT TTTGTCGGAC 551 GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAGGATTCCCGCCTTTAAG 601 CAGCCTGCTC CAAGCAAAAT CGGGCAGGGC AGTGTTCAAA ATATCGTCCT651 GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAGCTG TTTGGCTACG 701GACGCGAAAC TTCGCCGTTT TTAACCCGGC TGTCGCAAGC CGATTTTAAG 751 CCGATTGTGAAACAAAGTTA TTCCGCAGGC TTTATGACTG CAGTGTCCCT 801 GCCCAGTTTT TTCAATGCGATACCGCACGC CAACGGCTTG GAACAAATCA 851 GCGGCGGCGA TACCAATATG TTCCGCCTCGCCAAAGAGCA GGGCTATGAA 901 ACGTATTTTT ACAGCGCGCA GGCGGAAAAC GAGATGGCGATTTTGAACTT 951 AATCGGTAAG AAATGGATAG ACCATCTGAT TCAGCCGACG CAACTTGGCT1001 ACGGCAACGG CGACAATATG CCCGATGAGA AGCTGCTGCC GTTGTTCGAC 1051AAAATCAATT TGCAGCAGGG CAAGCATTTT ATCGTGTTGC ACCAACGCGG 1101 TTCGCACGCCCCATACGGCG CATTGTTGCA GCCTCAAGAT AAAGTATTCG 1151 GCGAAGCCGA TATTGTGGATAAGTACGACA ACACCATCCA CAAAACCGAC 1201 CAAATGATTC AAACCGTATT CGAGCAGCTGCAAAAGCAGC CTGACGGCAA 1251 CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTTCGCCAAGATA 1301 TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATCTCGT GCCGCTAGTG1351 TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC AGGCTTTTGC 1401GCCTTGCGAG ATTGCCTTCC ATCAGCAGCT TTCAACGTTC CTGATTCACA 1451 CGTTGGGCTACGATATGCCG GTTTCAGGTT GTCGCGAAGG CTCGGTAACG 1501 GGCAACCTGA TTACGGGTGATGCAGGCAGC TTGAACATTC GCGACGGCAA 1551 GGCGGAATAT GTTTATCCGC AATGA

This corresponds to the amino acid sequence <SEQ ID 306; ORF81-1>:

1 MKKSFLTLVL YSSLLTASEI AYRFVFGIET LPAAKIAETF ALTFVIAALY 51 LFARYKVTRLLIAVFFAFSI IANNVHYAVY QSWMTGINYW LMLKEVTEVG 101 SAGASMLDKL WLPVLWGVLEVMLFCSLAKF RRKTHFSADI LFAFLMLMIF 151 VRSFDTKQEH GISPKPTYSR IKANYFSFGYFVGRVLPYQL FDLSRIPAFK 201 QPAPSKIGQG SVQNIVLIMG ESESAAHLKL FGYGRETSPFLTRLSQADFK 251 PIVKQSYSAG FMTAVSLPSF FNAIPHANGL EQISGGDTNM FRLAKEQGYE301 TYFYSAQAEN EMAILNLIGK KWIDHLIQPT QLGYGNGDNM PDEKLLPLFD 351KINLQQGKHF IVLHQRGSHA PYGALLQPQD KVFGEADIVD KYDNTIHKTD 401 QMIQTVFEQLQKQPDGNWLF AYTSDHGQYV RQDIYNQGTV QPDSYLVPLV 451 LYSPDKAVQQ AANQAFAPCEIAFHQQLSTF LIHTLGYDMP VSGCREGSVT 501 GNLITGDAGS LNIRDGKAEY VYPQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF81 shows 84.7% identity over a 85aa overlap and 99.2% identity over a121aa overlap with an ORF (ORF81a) from strain A of N. meningitidis:

The complete length ORF81a nucleotide sequence <SEQ ID 307> is:

1 ATGAAAAAAT CCCTTTTCGT TCTCTTTCTG TATTCGTCCC TACTTACTGC 51 CAGCGAAATTGCTTATCGCT TTGTATTCGG AATTGAAACC TTACCGGCTG 101 CAAAAATGGC AGAAACGTTTGCGCTGACAT TTGTGATTGC TGCGCTGTAT 151 CTGTTTGCGC GTTATAAGGC AACGCGTTTGTTGATTGCGG TGTTTTTCGC 201 GTTCAGCATT ATTGCCAACA ATGTGCATTA CGCGGTTTATCAAAGCTGGA 251 TAACGGGCAT TAATTATTGG CTGATGCTGA AAGAGATTAC CGAAGTTGGC301 GGCGCAGGGG CGTCGATGTT GGATAAGTTG TGGCTGCCTG CGTTGTGGGG 351CGTGTTGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC CGCCGTAAGA 401 CGCATTTTTCTGCCGATATA CTGTTTGCCT TCCTAATGCT GATGATTTTC 451 GTGCGTTCGT TCGACACGAAACAAGAACAC GGTATTTCGC CCAAACCGAC 501 ATACAGCCGC ATCAAAGCCA ATTATTTCAGCTTCGGTTAT TTTGTCGGAC 551 GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAAGATTCCTGTGTTCAAA 601 CAGCCTGCTC CAAGCAGAAT CGGGCAAGGC AGTATTCAAA ATATCGTCCT651 GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAATTG TTTGGCTACG 701GGCGCGAAAC TTCGCCGTTT TTGACCCAGC TTTCGCAAGC CGATTTTAAG 751 CCGATTGTGAAACAAAGTTA TTCCGCAGGC TTTATGACGG CAGTATCCCT 801 GCCCAGTTTC TTTAACGTCATACCGCATGC CAACGGCTTG GAACAAATCA 851 GCGGCGGCGA TATTGTGGAT AAGTACGACAACACCATCCA CAAAACCGAC 901 CAAATGATTC AAACCGTATT CGAGCAGCTG CAAAAGCAGCCTGACGGCAA 951 CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTT CGCCAAGATA1001 TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATCTCGT GCCGCTGGTG 1051TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC AGGCTTTTGC 1101 GCCTTGCGAGATTGCCTTCC ATCAGCAGCT TTCAACGTTC CTGATTCACA 1151 CGTTGGGCTA CGATATGCCGGTTTCAGGTT GTCGCGAAGG CTCGGTAACG 1201 GGCAACCTGA TTACGGGTGA TGCAGGCAGCTTGAACATTC GCGACGGCAA 1251 GGCGGAATAT GTTTATCCGC AATGA

This encodes a protein having amino acid sequence <SEQ ID 308>:

1 MKKSLFVLFL YSSLLTASEI AYRFVFGIET LPAAKMAETF ALTFVIAALY 51LFARYKATRL LIAVFFAFSI IANNVHYAVY QSWITGINYW LMLKEITEVG 101GAGASMLDKL WLPALWGVLE VMLFCSLAKF RRKTHFSADI LFAFLMLMIF 151VRSFDTKQEH GISPKPTYSR IKANYFSFGY FVGRVLPYQL FDLSKIPVFK 201QPAPSRIGQG SIQNIVLIMG ESESAAHLKL FGYGRETSPF LTQLSQADFK 251PIVKQSYSAG FMTAVSLPSF FNVIPHANGL EQISGGDIVD KYDNTIHKTD 301QMIQTVFEQL QKQPDGNWLF AYTSDHGQYV RQDIYNQGTV QPDSYLVPLV 351LYSPDKAVQQ AANQAFAPCE IAFHQQLSTF LIHTLGYDMP VSGCREGSVT 401GNLITGDAGS LNIRDGKAEY VYPQ*

ORF81a and ORF81-1 show 77.9% identity in 524 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

The aligned aa sequences of ORF81 and a predicted ORF (ORF81.ng) from N.gonorrhoeae of the N- and C-termini show 82.4% and 97.5% identity in 85and 121 overlap, respectively:

The complete length ORF81ng nucleotide sequence <SEQ ID 309> is:

1 ATGAAAAAAT CCCTTTTCGT TCTCTTTCTG TATTCATCCC TACTTACCGC 51CAGCGAAATC GCCTATCGCT TTGTATTCGG AATTGAAACC TTACCGGCTG 101CAAAAATGGC GGAAACGTTT GCGCTGACAT TTATGATTGC TGCGCTGTAT 151CTGTTTGCGC GTTATAAGGC TTCGCGGCTG CTGATTGCGG TGTTTTTCGC 201GTTCAGCATG ATTGCCAACA ATGTGCATTA CGCGGTTTAT CAAAGCTGGA 251TGACGGGTAT TAACTATTGG CTGATGCTGA AAGAGGTTAC CGAAGTCGGC 301AGCGCGGGCG CGTCGATGTT GGATAAGTTG TGGCTGCCTG CTTTGTGGGG 351CGTGGCGGAA GTCATGTTGT TTTGCAGCCT TGCCAAGTTC CGCCGTAAGA 401CGCATTTTTC TGCCGATATA CTGTTTGCCT TCCTAATGCT GATGATTTTC 451GTGCGTTCGT TCGACACGAA ACAAGAGCAC GGTATTTCGC CCAAACCGAC 501ATACAGCCGC ATCAAAGCCA ATTATTTCAG CTTCGGTTAT TTTGTCGGGC 551GCGTGTTGCC GTATCAGTTG TTTGATTTAA GCAAGATCCC TGTGTTCAAA 601CAGCCTGCTC CAAGCAAAAT CGGGCAAGGC AGTATTCAAA ATATCGTCCT 651GATTATGGGC GAAAGCGAAA GCGCGGCGCA TTTGAAATTG TTTGGTTACG 701GGCGCGAAAC TTCGCCGTTT TTAACCCGGC TGTCGCAAGC CGATTTTAAG 751CCGATTGTGA AACAAAGTTA TTCCGCAGGC TTTATGACGG CAGTATCCCT 801GCCCAGTTTC TTTAACGTCA TACCGCACGC CAACGGCTTG GAACAAATCA 851GCGGCGGCGA TACCAATATG TTCCGCCTCG CCAAAGAGCA GGGCTATGAA 901ACGTATTTTT ACAGTGCCCA GGCTGAAAAC CAAATGGCAA TTTTGAACTT 951AATCGGTAAG AAATGGATAG ACCATCTGAT TCAGCCGACG CAACTTGGCT 1001ACGGCAACGG CGACAATATG CCCGATGAGA AGCTGCTGCC GTTGTTCGAC 1051AAAATCAATT TGCAGCAGGG CAGGCATTTT ATCGTGTTGC ACCAACGCGG 1101TTCGCACGCC CCATACGGCG CATTGTTGCA GCCTCAAGAT AAAGTATTCG 1151GCGAAGCCGA TATTGTGGAT AAGTACGACA ACACCATCCA CAAAACCGAC 1201CAAATGATTC AAACCGTATT CGAGCAGCTG CAAAAGCAGC CTGACGGCAA 1251CTGGCTGTTT GCCTATACCT CCGATCATGG CCAGTATGTG CGCCAAGATA 1301TCTACAATCA AGGCACGGTG CAGCCCGACA GCTATATTGT GCCTCTGGTT 1351TTGTACAGCC CGGATAAGGC CGTGCAACAG GCTGCCAACC AGGCTTTTGC 1401GCCTTGCGAG ATTGCCTTCC ATCAGCAGCT TTCAACGTTC CTGATTCACA 1451CGTTGGGCTA CGATATGCCG GTTTCAGGTT GTCGCGAAGG CTCGGTAACA 1501GGCAACCTGA TTACGGGCGA TGCAGGCAGC TTGAACATTC GCAACGGCAA 1551GGCGGAATAT GTTTATCCGC AATAA

This encodes a protein having amino acid sequence <SEQ ID 310>:

1 MKKSLFVLFL YSSLLTASEI AYRFVFGIET LPAAKMAETF ALTFMIAALY 51LFARYKASRL LIAVFFAFSM IANNVHYAVY QSWMTGINYW LMLKEVTEVG 101SAGASMLDKL WLPALWGVAE VMLFCSLAKF RRKTHFSADI LFAFLMLMIF 151VRSFDTKQEH GISPKPTYSR IKANYFSFGY FVGRVLPYQL FDLSKIPVFK 201QPAPSKIGQG SIQNIVLIMG ESESAAHLKL FGYGRETSPF LTRLSQADFK 251PIVKQSYSAG FMTAVSLPSF FNVIPHANGL EQISGGDTNM FRLAKEQGYE 301TYFYSAQAEN QMAILNLIGK KWIDHLIQPT QLGYGNGDNM PDEKLLPLFD 351KINLQQGRHF IVLHQRGSHA PYGALLQPQD KVFGEADIVD KYDNTIHKTD 401QMIQTVFEQL QKQPDGNWLF AYTSDHGQYV RQDIYNQGTV QPDSYIVPLV 451LYSPDKAVQQ AANQAFAPCE IAFHQQLSTF LIHTLGYDMP VSGCREGSVT 501GNLITGDAGS LNIRNGKAEY VYPQ*

ORF81ng and ORF81-1 show 96.4% identity in 524 aa overlap:

Furthermore, ORF81ng shows significant homology to an E. coli OMP:

gi|1256380 (U50906) outer membrane adherence protein-associatedprotein [E. coli] Length = 547 Score = 87.4 bits (213), Expect = 2e−16Identities = 122/468 (26%), Positives = 198/468 (42%), Gaps =70/468 (14%) Query: 25VFGIETLPAAKMAETFA-LTFMIAALYLFARYKAS--RLLIAVFFAFSMIANNVHYAVYQ 81VFGI  L A+  A     L F +  + +  R  +   RLL+A  F   + A ++  ++Y Sbjct: 29VFGITNLVASSGAHMVQRLLFFVLTILVVKRISSLPLRLLVAAPFVL-LTAADMSISLY- 86 Query:82 SWMT-------GINYWLMLKEVTEVGSAGASMLDKLWLPALWGVAEVMLFCSLAKFRRKT 134SW T       G    ++  +  EV    A ML  ++ P L   A + L       + Sbjct: 87SWCTFGTTFNDGFAISVLQSDPDEV----AKMLG-MYSPYLCAFAFLSLLFLAVIIKYDV 141 Query:135 HFSADILFAFLMLMIFVRSF---------DTKQEHGISPKPTYSRIKAN--YFSFGYFVG 183      +   L+L++   S          D K ++  SP    SR      +F+  YF Sbjct: 142SLPTKKVTGILLLIVISGSLFSACQFAYKDAKNKNAFSPYILASRFATYTPFFNLNYFAL 201 Query:184 RVLPYQ--LFDLSKIPVFKQPAPSKIGQGSIQNIVLIMGESESAAHLKLFGYGRETSPFL 241    +Q  L   + +P F+      +    I   VLI+GES    ++ L+GY R T+P + Sbjct: 202AAKEHQRLLSIANTVPYFQL----SVRDTGIDTYVLIVGESVRVDNMSLYGYTRSTTPQV 257 Query:242 TRLSQADFKPIVKQSYSAGFMTAVSLP---SFFNVIPHANGLEQISGGDTNMFRLAKEQG 298   +Q     +  Q+ S    TA+S+P   +  +V+ H      I     N+  +A + G Sbjct: 258E--AQRKQIKLFNQAISGAPYTALSVPLSLTADSVLSH-----DIHNYPDNIINMANQAG 310 Query:299 YETYFYSAQA---ENQMAILNLIGKKWIDHLIQPTQLGYGNGDNMPDEKLLPLFDKINLQ 355++T++ S+Q+   +N  A+ ++         ++  +  Y  G    DE LLP   +   Q Sbjct: 311FQTFWLSSQSAFRQNGTAVTSI--------AMRAMETVYVRGF---DELLLPHLSQALQQ 359 Query:356 --QGRHFIVLHQRGSHAPYGALLQPQDKVFGEADIVDK-YDNTIHKTDQMIQTVFEQLQK 412  Q +  IVLH  GSH P  +       VF   D  D  YDN+IH TD ++  VFE L+ Sbjct: 360NTQQKKLIVLHLNGSHEPACSAYPQSSAVFQPQDDQDACYDNSIHYTDSLLGQVFELLK- 418 Query:413 QPDGNWLFAYTSDHG---QYVRQDIYNQG--TVQPDSYIVPL-VLYSP 454  D      Y +DHG      ++++Y  G       +Y VP+ + YSP Sbjct: 419--DRRASVMYFADHGLERDPTKKNVYFHGGREASQQAYHVPMFIWYSP 464

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 37

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 311>:

1 ...ACCCTGCTCC TCTTCATCCC CCTCGTCCTC ACAC.GTGCG GCACACTGAC 51   CGGCATACTC GCCCaCGGCG GCGGCAAACG CTTTGCCGTC GAACAAGAAC 101   TCGTCGCCGC ATCGTCCCGC GCCGCCGTCA AAGAAATGGA TTTGTCCGCC 151   yTAAAAGGAC GCAAAGCCGC CyTTTACGTC TCCGTTATGG GCGACCAAGG 201   TTCGGGCAAC ATAAGCGGCG GACGCTACTC TATCGACGCA CTGATACGCG 251   GCGGCTACCA CAACAACCCC GAAAGTGCCA CCCAATACAG CTACCCCGCC 301   TACGACACTA CCGCCACCAC CAAATCCGAC GCGCTCTCCA GCGTAACCAC 351   TTCCACATCG CTTTTGAACG CCCCCGCCGC CGyCyTGACG AAAAACAGCG 401   GACGCAAAGG CGAACGcTCC GCCGGACTGT CCGTCAACGG CACGGGCGAC 451   TACCGCAACG AAACCCTGCT CGCCAACCCC CGCGACGTTT CCTTCCTGAC 501   CAACCTCATC CAAACCGTCT TCTACCTGCG CGGCATCGAA GTCgTACCGC 551   CCGrATACGC CGACACCGAC GTATTCGTAA CCGTCGACGT A...

This corresponds to the amino acid sequence <SEQ ID 312; ORF83>:

1 ..TLLLFIPLVL TXCGTLTGIL AHGGGKRFAV EQELVAASSR AAVKEMDLSA 51  LKGRKAAXYV SVMGDQGSGN ISGGRYSIDA LIRGGYHNNP ESATQYSYPA 101  YDTTATTKSD ALSSVTTSTS LLNAPAAXLT KNSGRKGERS AGLSVNGTGD 151  YRNETLLANP RDVSFLTNLI QTVFYLRGIE VVPPXYADTD VFVTVDV..

Further work revealed the complete nucleotide sequence <SEQ ID 313>:

1 ATGAAAACCC TGCTCCTCCT CATCCCCCTC GTCCTCACAG CCTGCGGCAC 51ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT GCCGTCGAAC 101AAGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA AATGGATTTG 151TCCGCCCTAA AAGGACGCAA AGCCGCCCTT TACGTCTCCG TTATGGGCGA 201CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCTATC GACGCACTGA 251TACGCGGCGG CTACCACAAC AACCCCGAAA GTGCCACCCA ATACAGCTAC 301CCCGCCTACG ACACTACCGC CACCACCAAA TCCGACGCGC TCTCCAGCGT 351AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC CTGACGAAAA 401ACAGCGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT CAACGGCACG 451GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG ACGTTTCCTT 501CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC ATCGAAGTCG 551TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT CGACGTATTC 601GGCACCGTCC GCAGCCGTAC CGAACTGCAC CTCTACAACG CCGAAACCCT 651TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTTGACCGC GACAGCCGGA 701AACTGCTGAT TACCCCTAAA ACCGCCGCCT ACGAATCCCA ATACCAAGAA 751CAATACGCCC TTTGGACCGG CCCTTACAAA GTCAGCAAAA CCGTCAAAGC 801CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATTACCCCC TACGGCGACA 851CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG TAAAAAACCC 901GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT AA

This corresponds to the amino acid sequence <SEQ ID 314; ORF83-1>:

1 MKTLLLLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS SRAAVKEMDL 51SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN NPESATQYSY 101PAYDTTATTK SDALSSVTTS TSLLNAPAAA LTKNSGRKGE RSAGLSVNGT 151GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD TDVFVTVDVF 201GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLITPK TAAYESQYQE 251QYALWTGPYK VSKTVKASDR LMVDFSDITP YGDTTAQNRP DFKQNNGKKP 301DVGNEVIRRR KGG*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF83 shows 96.4% identity over a 197aa overlap with an ORF (ORF83a)from strain A of N. meningitidis:

The complete length ORF83a nucleotide sequence <SEQ ID 315> is:

1 ATGAAAACCC TGCTCNTCCT CATCCCCCTC GTCCTCACAG CCTGCGGCAC 51ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT GCCGTCGAAC 101AAGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA AATGGACTTG 151TCCGCCCTGA AAGGACGCAA AGCCGCCCTT TACGTCTCCG TTATGGGCGA 201CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCTATC GACGCACTGA 251TACGCGGCGG CTACCACAAC AACCCCGAAA GTGCCACCCA ATACAGCTAC 301CCCGCCTACG ACACTACCGC CACCACCAAA TCCGACGCGC TCTCCAGCGT 351AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC CTGACGAAAA 401ACAGCGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT CAACGGCACG 451GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG ACGTTTCCTT 501CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC ATCGAAGTCG 551TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT CGACGTATTC 601GGCACCGTCC GCAGCCGCAC CGAACTGCAC CTCTACAACG CCGAAACCCT 651TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTTGACCGC GACAGCCGGA 701AACTGCTGAT TGCCCCTAAA ACCGCCGCCT ACGAATCCCA ATACCAAGAA 751CAATACGCCC TCTGGATGGG ACCTTACAGC GTCGGCAAAA CCGTCAAAGC 801CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATCACCCCC TACGGCGACA 851CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG TAAAAAACCC 901GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT AA

This encodes a protein having amino acid sequence <SEQ ID 316>:

1 MKTLLXLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS SRAAVKEMDL 51SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN NPESATQYSY 101PAYDTTATTK SDALSSVTTS TSLLNAPAAA LTKNSGRKGE RSAGLSVNGT 151GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD TDVFVTVDVF 201GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLIAPK TAAYESQYQE 251QYALWMGPYS VGKTVKASDR LMVDFSDITP YGDTTAQNRP DFKQNNGKKP 301DVGNEVIRRR KGG*

ORF83a and ORF83-1 show 98.4% identity in 313 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF83 shows 94.9% identity over a 197aa overlap with a predicted ORF(ORF83.ng) from N. gonorrhoeae:

The complete length ORF83ng nucleotide sequence <SEQ ID 317> is:

1 ATGAAAACCC TGCTCCTCCT CATCCCCCTC GTACTCACCG CCTGCGGCAC 51ACTGACCGGC ATACCCGCCC ACGGCGGCGG CAAACGCTTT GCCGTCGAAC 101AGGAACTCGT CGCCGCATCG TCCCGCGCCG CCGTCAAAGA AATGGACTTG 151TCCGCCCTGA AAGGACGCAA AGCCGCCCTT TACGTCTCCG TTATGGGCGA 201CCAAGGTTCG GGCAACATAA GCGGCGGACG CTACTCCATC GACGCACTGA 251TACGCGGCGG CTACCACAAC AACCCCGACA GCGCCACCCG ATACAGCTAC 301CCCGCCTATG ACACTACCGC CACCACCAAA TCCGACGCGC TCTCCGGCGT 351AACCACTTCC ACATCGCTTT TGAACGCCCC CGCCGCCGCC CTGACGAAAA 401ACAACGGACG CAAAGGCGAA CGCTCCGCCG GACTGTCCGT CAACGGCACG 451GGCGACTACC GCAACGAAAC CCTGCTCGCC AACCCCCGCG ACGTTTCCTT 501CCTGACCAAC CTCATCCAAA CCGTCTTCTA CCTGCGCGGC ATCGAAGTCG 551TACCGCCCGA ATACGCCGAC ACCGACGTAT TCGTAACCGT CGACGTATTC 601GGCACCGTCC GCAGCCGTAC CGAACTGCAC CTCTACAACG CCGAAACCCT 651TAAAGCCCAA ACCAAGCTCG AATATTTCGC CGTCGACCGC GACAGCCGGA 701AACTGCTGAT TGCCCCTAAA ACCGCCGCCT ACGAATCCCA ATACCAAGAA 751CAATACGCCC TCTGGATGGG ACCTTACAGC GTCGGCAAAA CCGTCAAAGC 801CTCAGACCGC CTGATGGTCG ATTTCTCCGA CATCACCCCC TACGGCGACA 851CAACCGCCCA AAACCGTCCC GACTTCAAAC AAAACAACGG TAAAAACCCC 901GATGTCGGCA ACGAAGTCAT CCGCCGCCGC AAAGGAGGAT AA

This encodes a protein having amino acid sequence <SEQ ID 318>:

1 MKTLLLLIPL VLTACGTLTG IPAHGGGKRF AVEQELVAAS SRAAVKEMDL 51SALKGRKAAL YVSVMGDQGS GNISGGRYSI DALIRGGYHN NPDSATRYSY 101PAYDTTATTK SDALSGVTTS TSLLNAPAAA LTKNNGRKGE RSAGLSVNGT 151GDYRNETLLA NPRDVSFLTN LIQTVFYLRG IEVVPPEYAD TDVFVTVDVF 201GTVRSRTELH LYNAETLKAQ TKLEYFAVDR DSRKLLIAPK TAAYESQYQE 251QYALWMGPYS VGKTVKASDR LMVDFSDITP YGDTTAQNRP DFKQNNGKNP 301DVGNEVIRRR KGG*

ORF83ng and ORF83-1 show 97.1% identity in 313 aa overlap

Based on this analysis, including the presence of a putativeATP/GTP-binding site motif A (P-loop) in the gonococcal protein(double-underlined) and a putative prokaryotic membrane lipoproteinlipid attachment site (single-underlined), it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 38

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 319>:

1 ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG GGAAAACATT 51AAAAATGGTT TCCATGATGG CGAATGATGA AATGTTTAAG CCTGATGAAA 101AAGCCATACG CCGTAAAGTA TTTACGAACA TAAAAGGCTT GAAAATACCG 151CACACCTACA TAGAAACGGA CGCAAAAAAG CTGCCGAAAT CGACAGATGA 201GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAG CCCGAAAATA 251TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG GCCGGCACGC 301TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA ATACGCACAG 351ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGTCCT AAGCTTCTAG 401ATCAAAATCT TAGAACGCTT GTACGGAAAC ATTACCACAT CGCTTCAAAC 451AAGATGGGTA TGCGTACGCT TTTAGAATGG AAAATATGCG CGGACGATCC 501CGTAAAAATG GCATCAAGCG CATTCTCCAG TATCTATACA CTGGATAAAA 551AAGTTTATGA CTTGTAysrr TmmGCGGAAG TTCATACCGT AAATAAGGTC 601AAGCGGTCAA AGTGGTTTTA CACTCTGCCa GTAATAGTAT TGCTGATTCC 651CGTGTTTGTC GGCCTGTCCT ATAAAATGTT GagCaGTTAC GGAAAAAAAC 701aGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA GCAGGCAGTA 751CTTCCGGATA AAACAGAAGG CGAGCCGGTA AATAACGGCA ACCTTACCGC 801AGATATGTTT GTTCCGACAT TGTCCGAaAA ACCCGrAAGC AAGCcgaTTT 851ATAACGGTGT AAGGCAGGTA AGAACCTTTG AATATATAGC AGGCTGTATA 901GAAGGCGGAA GAACCGGATG CGCCTGCTAT TCGCaTCAAG GGACGGCATt 951gaAAGAAGTG ACGGaGTTGA TGTGccaAgG aCTATGTaAA AAacGGCTTG 1001CCGTTTAACC CaTACAAAGA AGAAAGCCAA GGGCAGGAAG TTCAGCAAAG 1051CGCGCAgCAA CATTCGGACA GGGCGcCAAG TTGCCACATT GGGCGGAAAA 1101CCGTAGCAGA ACCTAATGTA CGATAATTGG GAAGAACGCG GGAAACCGTT 1151TGAAGGAATC GGaCGQGGGC GTGGTCGGAT CGGCAAACTG A

This corresponds to the amino acid sequence <SEQ ID 320; ORF84>:

1 MAEICLITGT PGSGKTLKMV SMMANDEMFK PDEKAIRRKV FTNIKGLKIP 51HTYIETDAKK LPKSTDEQLS AHDMYEWIKK PENIGSIVIV DEAQDVWPAR 101SAGSKIPENV QWLNTHRHQG IDIFVLTQGP KLLDQNLRTL VRKHYHIASN 151KMGMRTLLEW KICADDPVKM ASSAFSSIYT LDKKVYDLYX XAEVHTVNKV 201KRSKWFYTLP VIVLLIPVFV GLSYKMLSSY GKKQEEPAAQ ESAATEQQAV 251LPDKTEGEPV NNGNLTADMF VPTLSEKPXS KPIYNGVRQV RTFEYIAGCI 301EGGRTGCACY SHQGTALKEV TELMCKDYVK NGLPFNPYKE ESQGQEVQQS 351AQQHSDRAQV ATLGGKPXQN LMYDNWEERG KPFEGIGGGV VGSAN*

Further work revealed the complete nucleotide sequence <SEQ ID 321>:

1 ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG GGAAAACATT 51AAAAATGGTT TCCATGATGG CGAATGATGA AATGTTTAAG CCTGATGAAA 101ACGGCATACG CCGTAAAGTA TTTACGAACA TAAAAGGCTT GAAAATACCG 151CACACCTACA TAGAAACGGA CGCAAAAAAG CTGCCGAAAT CGACAGATGA 201GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAG CCCGAAAATA 251TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG GCCGGCACGC 301TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA ATACGCACAG 351ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGTCCT AAGCTTCTAG 401ATCAAAATCT TAGAACGCTT GTACGGAAAC ATTACCACAT CGCTTCAAAC 451AAGATGGGTA TGCGTACGCT TTTAGAATGG AAAATATGCG CGGACGATCC 501CGTAAAAATG GCATCAAGCG CATTCTCCAG TATCTATACA CTGGATAAAA 551AAGTTTATGA CTTGTACGAA TCAGCGGAAG TTCATACCGT AAATAAGGTC 601AAGCGGTCAA AGTGGTTTTA CACTCTGCCA GTAATAGTAT TGCTGATTCC 651CGTGTTTGTC GGCCTGTCCT ATAAAATGTT GAGCAGTTAC GGAAAAAAAC 701AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA GCAGGCAGTA 751CTTCCGGATA AAACAGAAGG CGAGCCGGTA AATAACGGCA ACCTTACCGC 801AGATATGTTT GTTCCGACAT TGTCCGAAAA ACCCGAAAGC AAGCCGATTT 851ATAACGGTGT AAGGCAGGTA AGAACCTTTG AATATATAGC AGGCTGTATA 901GAAGGCGGAA GAACCGGATG CGCCTGCTAT TCGCATCAAG GGACGGCATT 951GAAAGAAGTG ACGGAGTTGA TGTGCAAGGA CTATGTAAAA AACGGCTTGC 1001CGTTTAACCC ATACAAAGAA GAAAGCCAAG GGCAGGAAGT TCAGCAAAGC 1051GCGCAGCAAC ATTCGGACAG GGCGCAAGTT GCCACATTGG GCGGAAAACC 1101GTAGCAGAAC CTAATGTACG ATAATTGGGA AGAACGCGGG AAACCGTTTG 1151AAGGAATCGG CGGGGGCGTG GTCGGATCGG CAAACTGA

This corresponds to the amino acid sequence <SEQ ID 322; ORF84-1>:

1 MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGIRRKV FTNIKGLKIP 51 HTYIETDAKKLPKSTDEQLS AHDMYEWIKK PENIGSIVIV DEAQDVWPAR 101 SAGSKIPENV QWLNTHRHQGIDIFVLTQGP KLLDQNLRTL VRKHYHIASN 151 KMGMRTLLEW KICADDPVKM ASSAFSSIYTLDKKVYDLYE SAEVHTVNKV 201 KRSKWFYTLP VIVLLIPVFV GLSYKMLSSY GKKQEEPAAQESAATEQQAV 251 LPDKTEGEPV NNGNLTADMF VPTLSEKPES KPIYNGVRQV RTFEYIAGCI301 EGGRTGCACY SHQGTALKEV TELMCKDYVK NGLPFNPYKE ESQGQEVQQS 351AQQHSDRAQV ATLGGKP*QN LMYDNWEERG KPFEGIGGGV VGSAN*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF84 shows 93.9% identity over a 395aa overlap with an ORF (ORF84a)from strain A of N. meningitidis:

The complete length ORF84a nucleotide sequence <SEQ ID 323> is:

1 ATGGCAGAGA TCTGTTTGAT AACCGGCACG CCCGGTTCAG GGAAAACATT 51 AAAAATGGTTTCCATGATGG CAAACGATGA AATGTTTAAG CCGGATGAAA 101 ACGGCATACG CCGTAAAGTATTTACGAACA TCAAAGGCTT GAAGATACCG 151 CACACCTACA TAGAAACGGA CGCGAAAAAGCTGCCGAAAT CGACAGATGA 201 GCAGCTTTCG GCGCATGATA TGTACGAATG GATAAAGAAGCCCGAAAATA 251 TCGGGTCTAT TGTCATTGTA GATGAAGCTC AAGACGTATG GCCGGCACGC301 TCGGCAGGTT CAAAAATCCC TGAAAATGTC CAATGGCTGA ATACGCACAG 351ACATCAGGGC ATTGATATAT TTGTTTTGAC TCAAGGCTCT AAGCTTCTAG 401 ATCAAAATCTTAGAACGCTT GTACGGAAAC ATTACCACAT CGCTTCAAAC 451 AAGATGGGTA TGCGTACGCTTTTAGAATGG AAAATATGCG CGGACGATCC 501 CGTAAAAATG GCATCAAGCG CATTCTCCAGTATCTATACA CTGGATAAAA 551 AAGTTTATGA CTTGTACGAA TCAGCGGAAG TTCATACCGTAAATAAGGTC 601 AAGCGGTCAA AATGGTTTTA TACTCTGCCA GTAATAATAT TGCTGATTCC651 CGTTTTTGTC GGCCTGTCCT ATAAAATGTT AAGTAGTTAT GGAAAAAAAC 701AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA TCAGGCAGTA 751 TTTCAGGATAAAACAGAAGG CGAGCCGGTA AACAACGGTA ACCTTACCGC 801 AGATATGTTT GTTCCGACATTGTCCGAAAA ACCCGAAAGC AAGCCGATTT 851 ATAACGGTGT AAGGCAGGTA AGAACCTTTGAATATATAGC AGGCTGTGTA 901 GAAGGCGGAA GAACCGGATG CACATGCTAT TCGCATCAAGGGACGGCATT 951 GAAAGAAATT ACAAAGGAAA TGTGCAAGGA TTACGCAAGA AACGGATTGC1001 CGTTTAACCC ATATAAAGAA GAAAGCCAAG GGCGGGATGT CCAGCAAAGT 1051GAGCAGCACC ATTCGGACAG ACCGCAAGTT GCCACGTTGG GCGGAAAGCC 1101 GTGGCAAAATCTTATGTATG ATAATTGGCA GGAGCGCGGA AAACCGTTTG 1151 AAGGAATCGG CGGGGGCGTGGTCGGATCGG CAAACTGA

This encodes a protein having amino acid sequence <SEQ ID 324>:

1 MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGIRRKV FTNIKGLKIP 51 HTYIETDAKKLPKSTDEQLS AHDMYEWIKK PENIGSIVIV DEAQDVWPAR 101 SAGSKIPENV QWLNTHRHQGIDIFVLTQGS KLLDQNLRTL VRKHYHIASN 151 KMGMRTLLEW KICADDPVKM ASSAFSSIYTLDKKVYDLYE SAEVHTVNKV 201 KRSKWFYTLP VIILLIPVFV GLSYKMLSSY GKKQEEPAAQESAATEHQAV 251 FQDKTEGEPV NNGNLTADMF VPTLSEKPES KPIYNGVRQV RTFEYIAGCV301 EGGRTGCTCY SHQGTALKEI TKEMCKDYAR NGLPFNPYKE ESQGRDVQQS 351EQHHSDRPQV ATLGGKPWQN LMYDNWQERG KPFEGIGGGV VGSAN*

ORF84a and ORF84-1 show 95.2% identity in 395 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF84 shows 94.2% identity over a 395aa overlap with a predicted ORF(ORF84.ng) from N. gonorrhoeae:

The complete length ORF84ng nucleotide sequence <SEQ ID 325> is:

1 ATGGCAGAAA TCTGTTTGAT AACCGGCACG CCCGGTTCAG GGAAAACATT 51 AAAAATGGTTTCCATGATGG CAAACGATGA AATGTTTAAG CCAGATGAAA 101 ACGGCGTACG CCGTAAAGTATTTACGAACA TCAAAGGTTT GAAGATACCG 151 CACACCCACA TAGAAACAGA CGCAAAGAAGCTGCCGAAAT CAACCGATGA 201 ACAGCTTTCG GCGCATGATA TGTATGAATG GATCAAGAAGCCTGAAAacg 251 tcggcgCAAT CGTTATTGTC GATGAGGCGC AAGACGTATG GCCCGCACGC301 TccgCAGGTT CGAAAATCCC CGAAAACGTC CAATGGCTGA ACACACACAG 351GCATCAGGGC ATAGATATAT TTGTATTGAC ACAAGGTCCT AAACTCTTAG 401 ATCAGAACTTGCGAACATTG GTTAAAAGAC ATTACCACAT TGCGGCCAAC 451 AAAATGGGTT TGCGTACCCTGCTTGAATGG AAAGTATGCG CGGATGACCC 501 GGTAAAAATG GCATCAAGTG CATTTTCCAGTATCTACACA CTGGATAAAA 551 AAGTTTATGA CTTGTACGAA TCCGCAGAAA TTCACACGGTAAACAAAGTC 601 AAGCGTTCAA AATGGTTTTA TGCATTGCCC GTCATCATAT TATTGATTCC651 GCTATTTGTC GGTTTGTCTT ACAAAATGTT GGGCAGTTAC GGAAAAAAAC 701AGGAAGAACC CGCAGCACAA GAATCGGCGG CAACAGAACA GCAGGCAGTA 751 CTTCCGGATAAAACAGAAGG AGAATCGGTG AATAACGGAA ACCTTACGGC 801 AGATATGTTT GTTCCGACATTGCCCGAAAA ACCCGAAAGC AAGCCGATTT 851 ATAACGGTGT AAGGCAGGTA AGGACCTTTGAATATATAGC AGGCTGTATA 901 GAAGGCGGAA GAACCGGATG CACCTGCTAT TCGCATCAAGGGACGGCATT 951 GAAAGAAGTG ACGGAGTTGA TGTGCAAGGA CTATGTAAAA AACGGCTTGC1001 CGTTTAACCC ATACAAAGAA GAAAGCCAAG GGCAGGAAGT TCAGCAAAGC 1051GCGCAGCAAC ATTCGGACAG GGCGCAAGTT GCCACCTTGG GCGGAAAACC 1101 GCAGCAGAACCTAATGTACG ACAATTGGGA AGAACGCGGG AAACCGTTTG 1151 AAGGAATCGG CGGGGGCGTGGTCGGATCGG CAAACTGA

This encodes a protein having amino acid sequence <SEQ ID 326>:

1 MAEICLITGT PGSGKTLKMV SMMANDEMFK PDENGVRRKV FTNIKGLKIP 51 HTHIETDAKKLPKSTDEQLS AHDMYEWIKK PENVGAIVIV DEAQDVWPAR 101 SAGSKIPENV QWLNTHRHQGIDIFVLTQGP KLLDQNLRTL VKRHYHIAAN 151 KMGLRTLLEW KVCADDPVKM ASSAFSSIYTLDKKVYDLYE SAEIHTVNKV 201 KRSKWFYALP VIILLIPLFV GLSYKMLGSY GKKQEEPAAQESAATEQQAV 251 LPDKTEGESV NNGNLTADMF VPTLPEKPES KPIYNGVRQV RTFEYIAGCI301 EGGRTGCTCY SHQGTALKEV TELMCKDYVK NGLPFNPYKE ESQGQEVQQS 351AQQHSDRAQV ATLGGKPQQN LMYDNWEERG KPFEGIGGGV VGSAN*

ORF84ng and ORF84-1 show 95.4% identity in 395 aa overlap:

Based on this analysis, includng the presence of a putativetransmembrane domain (single-underlined) in the gonococcal protein, anda putative ATP/GTP-binding site motif A (P-loop, double-underlined), itis predicted that the proteins from N. meningitidis and N. gonorrhoeae,and their epitopes, could be useful antigens for vaccines ordiagnostics, or for raising antibodies.

Example 39

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 327>:

1 GTGGTTTTCC TGAATGCCGA CAACGGGATA TTGGTTCAGG ACTTGCCTTT 51 TGAAGTCAAACTGAAAAAAT TCCATATCGA TTTTTACAAT ACGGGTATGC 101 CGCGTGATTT CGCCAGCGATATTGAAGTGA CGGACAAGGC AACCGGTGAG 151 AAACTCGAGC GCACCATCCG CGTGAACCATCCTTTGACCT TGCACGGCAT 201 CACGATTTAT CAGGCGAGTT TTGCCGACGG CGGTTCGGATTTGACATTCA 251 AGGCGTGGAA TTTGGGTGAT GCTTCGCGCG AGCCTGTCGT GTTGAAGGCA301 ACATCCATAC ACCAGTTTCC GTTGGAAATT GGCAAACACA AATATCGTCT 351TGAGTTCGAT CAGTTCACTT CTATGAATGT GGAGGACATG AGCGAGGGCG 401 CGGAACGGGAAAAAAGCCTG AAATCCACGC TGCCCGATGT CCGCGCCGTT 451 ACTCAGGAAG GTCACAAATACACCAAT... .......... .....TACCG 501 TATCCGTGAT GCGCCAGGCC AGGCGGTCGAATATAAAAAC TATATGCTGC 551 CGGTTTTGCA GGAACAGGAT TATTTTTGGA TTACCGGCACGCGCAGCGC. 601 TTGCAGCAGC AATACCGCTG GCTGCGTATC CCCTTGGACA AGCAGTTGAA651 AGCGGACACC TTTATGGCAT TGCGTGAGTT TTTGAAAGAT GGGGAAGGGC 701GCAAACGTCT .GTTGCCGAC GCAACCAAAG GCGCACCTGC CGAAATCCGC 751 GAACAATTCATGCTGGCTGC GGAAAACACG CTGAACATCT TTGCACAAAA 801 AGGCTATTTG GGATTGGACGAATTTATTAC GTCCAATATC CCGAAAGAGC 851 AGCAGGATAA GATGCAGGGC TATTTCTACGAAATGCTTTA CGGCGTGATG 901 AACGCTGCTT TGGATGAAAC CAT.ACCCGG TACGGCTTGCCCGAATGGCA 951 GCAGGATGAA GCGCGGAATC GTTTCCTGCT GCACAGTATG GATGCGTACA1001 CGGGTTTGAC CGAATATCCC GCGCCTATGC TGCTGCAACT TGATGGGTTT 1051TCCGAGGTGC GTTCGTCGGG TTTGCAGATG ACCCGTTCCC C.GGTCCGCT 1101 TTTGGTCTATCTC...

This corresponds to the amino acid sequence <SEQ ID 328; ORF88>:

1 MVFLNADNGI LVQDLPFEVK LKKFHIDFYN TGMPRDFASD IEVTDKATGE 51 KLERTIRVNHPLTLHGITIY QASFADGGSD LTFKAWNLGD ASREPVVLKA 101 TSIHQFPLEI GKHKYRLEFDQFTSMNVEDM SEGAEREKSL KSTLPDVRAV 151 TQEGHKYTNX XXXXXYRIRD APGQAVEYKNYMLPVLQEQD YFWITGTRSX 201 LQQQYRWLRI PLDKQLKADT FMALREFLKD GEGRKRXVADATKGAPAEIR 251 EQFMLAAENT LNIFAQKGYL GLDEFITSNI PKEQQDKMQG YFYEMLYGVM301 NAALDETXTR YGLPEWQQDE ARNRFLLHSM DAYTGLTEYP APMLLQLDGF 351SEVRSSGLQM TRSXGPLLVY L...

Further work revealed the complete nucleotide sequence <SEQ ID 329>:

1 ATGAGTAAAT CCCGTAGATC TCCCCCACTT CTTTCCCGTC CGTGGTTCGC 51 TTTTTTCAGCTCCATGCGCT TTGCAGTCGC TTTGCTCAGT CTGCTGGGTA 101 TTGCATCGGT TATCGGTACGGTGTTGCAGC AAAACCAGCC GCAGACGGAT 151 TATTTGGTCA AATTCGGATC GTTTTGGGCGCAGATTTTTG GTTTTCTGGG 201 ACTGTATGAC GTCTATGCTT CGGCATGGTT TGTCGTTATCATGATGTTTT 251 TGGTGGTTTC TACCAGTTTG TGCCTGATTC GCAATGTGCC GCCGTTCTGG301 CGCGAAATGA AGTCTTTTCG GGAAAAGGTT AAAGAAAAAT CTCTGGCGGC 351GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCGCCC GAGGTTGCCA 401 AACGTTATCTGGAAGTACAA GGTTTTCAGG GAAAAACCAT TAACCGTGAA 451 GACGGGTCGG TTCTGATTGCCGCCAAAAAA GGCACAATGA ACAAATGGGG 501 CTATATCTTT GCCCATGTTG CTTTGATTGTCATTTGCCTG GGCGGGTTGA 551 TAGACAGTAA CCTGCTGTTG AAACTGGGTA TGCTGACCGGTCGGATTGTT 601 CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG AAAGTATTTT651 GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT TCCGAGGGGC 701AGAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT ATTGGTTCAG 751 GACTTGCCTTTTGAAGTCAA ACTGAAAAAA TTCCATATCG ATTTTTACAA 801 TACGGGTATG CCGCGTGATTTCGCCAGCGA TATTGAAGTG ACGGACAAGG 851 CAACCGGTGA GAAACTCGAG CGCACCATCCGCGTGAACCA TCCTTTGACC 901 TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACGGCGGTTCGGA 951 TTTGACATTC AAGGCGTGGA ATTTGGGTGA TGCTTCGCGC GAGCCTGTCG1001 TGTTGAAGGC AACATCCATA CACCAGTTTC CGTTGGAAAT TGGCAAACAC 1051AAATATCGTC TTGAGTTCGA TCAGTTCACT TCTATGAATG TGGAGGACAT 1101 GAGCGAGGGCGCGGAACGGG AAAAAAGCCT GAAATCCACG CTGAACGATG 1151 TCCGCGCCGT TACTCAGGAAGGTAAAAAAT ACACCAATAT CGGCCCTTCC 1201 ATTGTTTACC GTATCCGTGA TGCGGCAGGGCAGGCGGTCG AATATAAAAA 1251 CTATATGCTG CCGGTTTTGC AGGAACAGGA TTATTTTTGGATTACCGGCA 1301 CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT CCCCTTGGAC1351 AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT TTTTGAAAGA 1401TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA GGCGCACCTG 1451 CCGAAATCCGCGAACAATTC ATGCTGGCTG CGGAAAACAC GCTGAACATC 1501 TTTGCACAAA AAGGCTATTTGGGATTGGAC GAATTTATTA CGTCCAATAT 1551 CCCGAAAGAG CAGCAGGATA AGATGCAGGGCTATTTCTAC GAAATGCTTT 1601 ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCGGTACGGCTTG 1651 CCCGAATGGC AGCAGGATGA AGCGCGGAAT CGTTTCCTGC TGCACAGTAT1701 GGATGCGTAC ACGGGTTTGA CCGAATATCC CGCGCCTATG CTGCTGCAAC 1751TTGATGGGTT TTCCGAGGTG CGTTCGTCGG GTTTGCAGAT GACCCGTTCC 1801 CCGGGTGCGCTTTTGGTCTA TCTCGGCTCG GTGCTGTTGG TATTGGGTAC 1851 GGTATTGATG TTTTATGTGCGCGAAAAACG GGCGTGGGTA TTGTTTTCAG 1901 ACGGCAAAAT CCGTTTTGCC ATGTCTTCGGCCCGCAGCGA ACGGGATTTG 1951 CAGAAGGAAT TTCCAAAACA CGTCGAGAGT CTGCAACGGCTCGGCAAGGA 2001 CTTGAATCAT GACTGA

This corresponds to the amino acid sequence <SEQ ID 330; ORF88-1>:

1 MSKSRRSPPL LSRPWFAFFS SMRFAVALLS LLGIASVIGT VLQQNQPQTD 51 YLVKFGSFWAQIFGFLGLYD VYASAWFVVI MMFLVVSTSL CLIRNVPPFW 101 REMKSFREKV KEKSLAAMRHSSLLDVKIAP EVAKRYLEVQ GFQGKTINRE 151 DGSVLIAAKKGTMNKWGYIF AHVALIVICL GGLIDSNLLL KLGMLTGRIV 201 PDNQAVYAKD FKPESILGASNLSFRGNVNI SEGQSADVVF LNADNGILVQ 251 DLPFEVKLKK FHIDFYNTGM PRDFASDIEVTDKATGEKLE RTIRVNHPLT 301 LHGITIYQAS FADGGSDLTF KAWNLGDASR EPVVLKATSIHQFPLEIGKH 351 KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE GKKYTNIGPS401 IVYRIRDAAG QAVEYKNYML PVLQEQDYFW ITGTRSGLQQ QYRWLRIPLD 451KQLKADTFMA LREFLKDGEG RKRLVADATK GAPAEIREQF MLAAENTLNI 501 FAQKGYLGLDEFITSNIPKE QQDKMQGYFY EMLYGVMNAA LDETIRRYGL 551 PEWQQDEARN RFLLHSMDAYTGLTEYPAPM LLQLDGFSEV RSSGLQMTRS 601 PGALLVYLGS VLLVLGTVLM FYVREKRAWVLFSDGKIRFA MSSARSERDL 651 QKEFPKHVES LQRLGKDLNH D*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF88 shows 95.7% identity over a 371aa overlap with an ORF (ORF88a)from strain A of N. meningitidis:

The complete length ORF88a nucleotide sequence <SEQ ID 331> is:

1 ATGAGTAAAT CCCGTAGATC TCCCCCACTT CTTTCCCGTC CGTGGTTCGC 51 TTTTTTCAGCTCCATGCGCT TTGCGGTCGC TTTGCTCAGT CTGCTGGGTA 101 TTGCATCGGT TATCGGTACGGTGTTGCAGC AAAACCAGCC GCAGACGGAT 151 TATTTGGTCA AATTCGGATC GTTTTGGGCGCAGATTTTTG GTTTTCTGGG 201 ACTGTATGAC GTCTATGCTT CGGCATGGTT TGTCGTTATCATGATGTTTT 251 TGGTGGTTTC TACCAGTTTG TGCCTGATTC GCAATGTGCC GCCGTTCTGG301 CGCGAAATGA AGTCTTTTCG GGAAAAGGTT AAAGAAAAAT CTCTGGCGGC 351GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCGCCC GAGGTTGCCA 401 AACGTTATCTGGAAGTACAA GGTTTTCAGG GAAAAACCAT TAACCGTGAA 451 GACGGGTCGG TTCTGATTGCCGCCAAAAAA GGCACAATGA ACAAATGGGG 501 CTATATCTTT GCCCATGTTG CTTTGATTGTCATTTGCCTG GGCGGGTTGA 551 TAGACAGTAA CCTGCTGTTG AAACTGGGTA TGCTGACCGGTCGGATTGTT 601 CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG AAAGTATTTT651 GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT TCCGAGGGGC 701AGAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT ATTGGTTCAG 751 GACTTGCCTTTTGAAGTCAA ACTGAAAAAA TTCCATATCG ATTTTTACAA 801 TACGGGTATG CCGCGCGATTTTGCCAGTGA TATTGAAGTA ACGGATAAGG 851 CAACCGGTGA GAAACTCGAG CGCACCATCCGCGTGAACCA TCCTTTGACC 901 TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACGGCGGTTCGGA 951 TTTGACATTC AAGGCGTGGA ATTTGGGTGA TGCTTCGCGC GAGCCTGTCG1001 TGTTGAAGGC AACATCCATA CACCAGTTTC CGTTGGAAAT TGGCAAACAC 1051AAATATCGTC TTGAGTTCGA TCAGTTTACT TCTATGAATG TGGAGGACAT 1101 GAGCGAGGGCGCGGAACGGG AAAAAAGCCT GAAATCCACG CTGAACGATG 1151 TCCGCGCCGT TACTCAGGAAGGTAAAAAAT ACACCAATAT CGGCCCTTCC 1201 ATTGTTTACC GTATCCGTGA TGCGGCAGGGCAGGCGGTCG AATATAAAAA 1251 CTATATGCTG CCGGTTTTGC AGGAACAGGA TTATTTTTGGATTACCGGCA 1301 CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT CCCCTTGGAC1351 AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT TTTTGAAAGA 1401TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA GGCGCACCTG 1451 CCGAAATCCGCGAACAATTC ATGCTGGCTG CGGAAAACAC GCTGAACATC 1501 TTTGCACAAA AAGGCTATTTGGGATTGGAC GAATTTATTA CGTCCAATAT 1551 CCCGAAAGAG CAGCAGGATA AGATGCAGGGCTATTTCTAC GAAATGCTTT 1601 ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCGGTACGGCTTG 1651 CCCGAATGGC AGCAGGATGA AGCGCGGAAT CGTTTCCTGC TGCACAGTAT1701 GGATGCGTAC ACGGGTTTGA CCGAATATCC CGCGCCTATG CTGCTGCAAC 1751TTGATGGGTT TTCCGAGGTG CGTTCGTCGG GTTTGCAGAT GACCCGTTCC 1801 CCGGGTGCGCTTTTGGTCTA TCTCGGCTCG GTGCTGTTGG TATTGGGTAC 1851 GGTATTGATG TTTTATGTGCGCGAAAAACG GGCGTGGGTA TTGTTTTCAG 1901 ACGGCAAAAT CCGTTTTGCC ATGTCTTCGGCCCGCAGCGA ACGGGATTTG 1951 CAGAAGGAAT TTCCAAAACA CGTCGAGAGT CTGCAACGGCTCGGCAAGGA 2001 CTTGAATCAT GACTGA

This encodes a protein having amino acid sequence <SEQ ID 332>:

1 MSKSRRSPPL LSRPWFAFFS SMRFAVALLS LLGIASVIGT VLQQNQPQTD 51 YLVKFGSFWAQIFGFLGLYD VYASAWFVVI MMFLVVSTSL CLIRNVPPFW 101 REMKSFREKV KEKSLAAMRHSSLLDVKIAP EVAKRYLEVQ GFQGKTINRE 151 DGSVLIAAKKGTMNKWGYIF AHVALIVICL GGLIDSNLLL KLGMLTGRIV 201 PDNQAVYAKD FKPESILGASNLSFRGNVNI SEGQSADVVF LNADNGILVQ 251 DLPFEVKLKK FHIDFYNTGM PRDFASDIEVTDKATGEKLE RTIRVNHPLT 301 LHGITIYQAS FADGGSDLTF KAWNLGDASR EPVVLKATSIHQFPLEIGKH 351 KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE GKKYTNIGPS401 IVYRIRDAAG QAVEYKNYML PVLQEQDYFW ITGTRSGLQQ QYRWLRIPLD 451KQLKADTFMA LREFLKDGEG RKRLVADATK GAPAEIREQF MLAAENTLNI 501 FAQKGYLGLDEFITSNIPKE QQDKMQGYFY EMLYGVMNAA LDETIRRYGL 551 PEWQQDEARN RFLLHSMDAYTGLTEYPAPM LLQLDGFSEV RSSGLQMTRS 601 PGALLVYLGS VLLVLGTVLM FYVREKRAWVLFSDGKIRFA MSSARSERDL 651 QKEFPKHVES LQRLGKDLNH D*

ORF88a and ORF88-1 100.0% identity in 671 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF88 shows 93.8% identity over a 371aa overlap with a predicted ORF(ORF88.ng) from N. gonorrhoeae:

An ORF88ng nucleotide sequence <SEQ ID 333> was predicted to encode aprotein having amino acid sequence <SEQ ID 334>:

1 MVFLNADNGM LVQDLPFEVK LKKFHIDFYN TGMPRDFASD IEVTDKATGE 51 KLERTIRVNHPLTLHGITIY QASFADGGSD LTFKAWNLRD ASREPVVLKA 101 TSIHQFPLEI GKHKYRLEFDQFTSMNVEDM SEGAEREKSL KSTLNDVRAV 151 TQEGKKYTNI GPSIVYRIRD AAGQAVEYKNYMLPILQDKD YFWLTGTRSG 201 LQQQYRWLRI PLDKQLKADT FMALREFLKD GEGRKRLVADATKDAPAEIR 251 EQFMLAAENT LNIFAQKGYL GLDEFITSNI PKGQQDKMQG YFYEMLYGVM301 NAALDETIRR YGLPEWQQDE ARNRFLLHSM DAYTGLTEYP APMLLQLDGF 351SEVRSSGLQM TRSPGALLVY LGSVLLVLGT VFMFYVPKKR AWVLFSNXKI 401 RFAMSSARSERDLQKEFPKH VESLQRLGKD LNHD*

Further work revealed the complete gonococcal DNA sequence <SEQ ID 335>:

1 ATGAGTAAAT CCCGTATATC TCCCACACTT CTTTCCCGTC CGTGGTTCGC 51 TTTTTTCAGCTCCATGCGCT TTGCGGTCGC TTTGCTCAGT CTGCTGGGTA 101 TTGCATCGGT TATCGGCACGGTGTTACAGC AAAACCAGCC GCAGACGGAT 151 TATTTGGTCA AATTCGGACC GTTTTGGACTCGGATTTTTG ATTTTTTGGG 201 TTTGTATGAT GTCTATGCTT CGGCATGGTT TGTCGTTATCATGATGTTTC 251 TGGTGGTTTC TACCAGTTTG TGTTTAATCC GTAACGTTCC GCCGTTTTGG301 CGCGAAATGA AGTCTTTCCG GGAAAAGGTT AAAGAAAAAT CTCTGGCGGC 351GATGCGCCAT TCTTCGCTGT TGGATGTAAA AATTGCCCCC GAAGTTGCCA 401 AACGTTATCTGGAGGTGCGG GGTTTTCAGG GAAAAACCGT CAGCCGTGAG 451 GACGGGTCGG TTCTGATTGCCGCCAAAAAA GGCAcaatga acaaATGGGG 501 CTATATCTTT GCccaagtag ctTTGATTGTCATTTGCCTG GGCGGGTTGA 551 TAGACAGTAA CCTGCTGCTG AAGCTGGGTA TGCTGGCCGGTCGGATTGTT 601 CCGGACAATC AGGCGGTTTA TGCCAAGGAT TTCAAGCCCG AAAGTATTTT651 GGGTGCGTCC AATCTCTCAT TTAGGGGCAA CGTCAATATT TCCGAGGGGC 701AAAGTGCGGA TGTGGTTTTC CTGAATGCCG ACAACGGGAT GTTGGTTCAG 751 GACTTGCCTTTTGAAGTCAA ACTGAAAAAA TTCCATATCG ATTTTTACAA 801 TACGGGTATG CCGCGCGATTTTGCCAGCGA TATTGAAGTA ACGGACAAGG 851 CAACCGGTGA GAAACTCGAG CGCACCATCCGCGTGAACCA TCCTTTGACC 901 TTGCACGGCA TCACGATTTA TCAGGCGAGT TTTGCCGACGGCGGTTCGGA 951 TTTGACATTC AAGGCGTGGA ATTTGAGGGA TGCTTCGCGC GAACCTGTCG1001 TGTTGAAGGC AACCTCCATA CACCAGTTTC CGTTGGAAAT CGGCAAACAC 1051AAATATCGTC TTGAGTTCGA TCAGTTCACT TCTATGAATG TGGAGGACAT 1101 GAGCGAGGGTGCGGAACGGG AAAAAAGCCT GAAATCCACT CTGAACGATG 1151 TCCGCGCCGT TACTCAGGAAGGTAAAAAAT ACACCAATAT CGGCCCTTCC 1201 ATCGTGTACC GCATCCGTGA TGcggCAGGGCAGGCGGTCG AATATAAAAA 1251 CTATATGCTG CCGATTTTGC AGGACAAAGA TTATTTTTGGCTGACCGGCA 1301 CGCGCAGCGG CTTGCAGCAG CAATACCGCT GGCTGCGTAT CCCCTTGGAC1351 AAGCAGTTGA AAGCGGACAC CTTTATGGCA TTGCGTGAGT TTTTGAAAGA 1401TGGGGAAGGG CGCAAACGTC TGGTTGCCGA CGCAACCAAA GACGCACCTG 1451 CCGAAATCCGCGAACAATTC ATGCTGGCTG CGGAAAACAC GCTGAATATC 1501 TTTGCGCAAA AAGGCTATTTGGGATTGGAC GAATTTATTA CGTCCAATAT 1551 CCCGAAAGGG CAGCAGGATA AGATGCAGGGCTATTTCTAC GAAATGCTTT 1601 ACGGCGTGAT GAACGCTGCT TTGGATGAAA CCATACGCCGGTACGGCTTG 1651 CCCGAATGGC AGCAGGATGA AGCGCGGAAC CGTTTCCTGC TGCACAGTAT1701 GGATGCCTAT ACGGGGCTGA CGGAATATCC CGCGCCTATG CTGCTCCAGC 1751TTGACGGGTT TTCCGAGGTG CGTTCCTCAG GTTTGCAGAT GACCCGTTCG 1801 CCGGGTGCGCTTTTGGTCTA TCtcggctcg gtattgttgg TTTTGGgtac 1851 ggtaTttatg tTTTATGTGCGCGAAAAACG GGCGTGGgta tTGTTTTCag 1901 aCGGCAAAAT CCGTTTTGCT ATGtCTTcggCCcgcagcga ACGGGATTTG 1951 cAGAaggaaT TTCCAAAACA CGtcgAGAGC CTGCAACggctcggcaaggA 2001 CttgaaTCAT GACTga

This corresponds to the amino acid sequence <SEQ ID 336; ORF88ng-1>:

1 MSKSRISPTL LSRPWFAFFS SMRFAVALLS LLGIASVIGT VLQQNQPQTD 51 YLVKFGPFWTRIFDFLGLYD VYASAWFVVI MMFLVVSTSL CLIRNVPPFW 101 REMKSFREKV KEKSLAAMRHSSLLDVKIAP EVAKRYLEVR GFQGKTVSRE 151 DGSVLIAAKKGTMNKWGYIF AQVALIVICL GGLIDSNLLL KLGMLAGRIV 201 PDNQAVYAKD FKPESILGASNLSFRGNVNI SEGQSADVVF LNADNGMLVQ 251 DLPFEVKLKK FHIDFYNTGM PRDFASDIEVTDKATGEKLE RTIRVNHPLT 301 LHGITIYQAS FADGGSDLTF KAWNLRDASR EPVVLKATSIHQFPLEIGKH 351 KYRLEFDQFT SMNVEDMSEG AEREKSLKST LNDVRAVTQE GKKYTNIGPS401 IVYRIRDAAG QAVEYKNYML PILQDKDYFW LTGTRSGLQQ QYRWLRIPLD 451KQLKADTFMA LREFLKDGEG RKRLVADATK DAPAEIREQF MLAAENTLNI 501 FAQKGYLGLDEFITSNIPKG QQDKMQGYFY EMLYGVMNAA LDETIRRYGL 551 PEWQQDEARN RFLLHSMDAYTGLTEYPAPM LLQLDGFSEV RSSGLQMTRS 601 PGALLVYLGS VLLVLGTVFM FYVREKRAWVLFSDGKIRFA MSSARSERDL 651 QKEFPKHVES LQRLGKDLNH D*

ORF88ng-1 and ORF88-1 show 97.0% identity in 671 aa overlap:

Furthermore, ORG88ng-1 shows homology with a hypothetical protein fromAquifex aeolicus:

gi|2984296 (AE000771) hypothetical protein [Aquifex aeolicus] Length= 537 Score = 94.4 bits (231), Expect = 2e−18. Identities = 91/334(27%), Positives = 159/334 (47%), Gaps = 59/334 (17%) Query: 16FAFFSSMRFAVALLSLLGIASVIG-TVLQQNQPQTDYLVKFGPFWTRIFDFLGLYDVYAS 74 + F+S++ A+ ++ +LGI S++G T ++QNQ    YL +FG         L L DV+ S Sbjct: 80YDFLASLKLAIFIMLVLGILSMLGSTYIKQNQSFEWYLDQFGYDVGIWIWKLWLNDVFHS 139 Query:75 AWFVVIMMFLVVSTSLCLIRNVPPFWREMKSFREKVKEKSLAAMRHSSLLDVKIAPEVAK 134 ++++ ++ L V+   C I+ +P  W++  S +E++ +    A +H   + VKI P+  K Sbjct: 140WYYILFIVLLAVNLIFCSIKRLPRVWKQAFS-KERILKLDEHAEKHLKPITVKI-PDKDK 197 Query:135 --RYLEVRGFQGKTVSREDGSVLIAAKKGTMNKWGYIFAQVALIVICLGGLIDSNLLLKL 192  ++L  +GF+   V  E   + + A+KG  ++ G     +AL+VI  G LID Sbjct: 198VLKFLLKKGFK-VFVEEEGNKLYVFAEKGRFSRLGVYITHIALLVIMAGALID------- 249 Query:193 GMLAGRIVPDNQAVYAKDFKPESILGASNLSFRGNVNISEGQSADVVFLNADNGMLVQDL 252                      +I+G      RG++ ++EG + DV+ + A+       L Sbjct: 250----------------------AIVGV-----RGSLIVAEGDTNDVMLVGAE--QKPYKL 280 Query:253 PFEVKLKKFHIDFY---NTGMPRDFA-------SDIEVTDKATGEKLER--TIRVNHPLT 300 PFV L  F I  Y   N  + + FA       SDIE+ +   G K+E   T++VN P Sbjct: 281PFAVHLIDFRIKTYAEENPNVDKRFAQAVSSYESDIEIIN---GGKVEAKGTVKVNEPFD 337 Query:301 LHGITIYQASFA--DGGSDLTFKAWNLRDASREP 332      ++QA++   DG S+     + + A  +P Sbjct: 338 FGRYRLFQATYGILDGTSGMGVIVVDRKKAHEDP 371

Based on this analysis, including the putative transmembrane domain inthe gonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 40

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 337>:

1 ATGATGAGTA ATAmAATGGm ACAAAAAGGG TTTACATTGA TTGmGmTGAT 51 GATAGTCGTCGCGATACTCG GCATTATCAG CGTCATTGCC ATACCTTCTT 101 ATCmAAGTTA TATTGAAAAAGGCTATCAGT CCCAGCTTTA TACGGAGATG 151 GyCGGTATCA ACAATATTTC CAAACAGTTTATTTTGAAAA ATCCCCTGGA 201 CGATAATCAG ACCATCGAGA ACAAACTGGA AATATTTGTCTCAGGCTATA 251 AGATGAATCC GAAAATTGCC AAAAAaTATA GTGTTTCGGT AAAGTTTGTC301 GATAAGGAAA AATCAAGGGC ATACAGGTTG GTCGGCGTTC CGAAGGCGGG 351GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC GACGGATACA 401 AATGCCGTGATGCCGCTTCT GCCCAAGCCC ATTTGGAGAC CTTGTCCTCA 451 GATGTCGGCT GTGAAGCCTTCTCTAATCGT AAAAAATAA

This corresponds to the amino acid sequence <SEQ ID 338; ORF89>:

1 MMSNXMXQKG FTLIXXMIVV AILGIISVIA IPSYXSYIEK GYQSQLYTEM 51 XGINNISKQFILKNPLDDNQ TIENKLEIFV SGYKMNPKIA KKYSVSVKFV 101 DKEKSRAYRL VGVPKAGTGYTLSVWMNSVG DGYKCRDAAS AQAHLETLSS 151 DVGCEAFSNR KK*

Further work revealed the complete nucleotide sequence <SEQ ID 339>:

1 ATGATGAGTA ATAAAATGGA ACAAAAAGGG TTTACATTGA TTGAGATGAT 51 GATAGTCGTCGCGATACTCG GCATTATCAG CGTCATTGCC ATACCTTCTT 101 ATCAAAGTTA TATTGAAAAAGGCTATCAGT CCCAGCTTTA TACGGAGATG 151 GTCGGTATCA ACAATATTTC CAAACAGTTTATTTTGAAAA ATCCCCTGGA 201 CGATAATCAG ACCATCGAGA ACAAACTGGA AATATTTGTCTCAGGCTATA 251 AGATGAATCC GAAAATTGCC AAAAAATATA GTGTTTCGGT AAAGTTTGTC301 GATAAGGAAA AATCAAGGGC ATACAGGTTG GTCGGCGTTC CGAAGGCGGG 351GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC GACGGATACA 401 AATGCCGTGATGCCGCTTCT GCCCAAGCCC ATTTGGAGAC CTTGTCCTCA 451 GATGTCGGCT GTGAAGCCTTCTCTAATCGT AAAAAATAA

This corresponds to the amino acid sequence <SEQ ID 340; ORF89-1>:

1 MMSNKMEQKG FTLIEMMIVV AILGIISVIA IPSYQSYIEK GYQSQLYTEM 51 VGINNISKQFILKNPLDDNQ TIENKLEIFV SGYKMNPKIA KKYSVSVKFV 101 DKEKSRAYRL VGVPKAGTGYTLSVWMNSVG DGYKCRDAAS AQAHLETLSS 151 DVGCEAFSNR KK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with PilE of N. gonorrhoeae (Accession Number Z69260).

ORF89 and PilE protein show 30% aa identity in 120a overlap:

orf89 8 QKGFTLIXXMIVVAILGIISVIAIPSYXSYIEKGYQSQLYTEMXGINNISKQFILKNPL- 66QKGFTLI  MIV+AI+GI++ +A+P+Y  Y  +   S+      G  +   ++ L + + PilE 5QKGFTLIELMIVIAIVGILAAVALPAYQDYTARAQVSEAILLAEGQKSAVTEYYLNHGIW 64 orf89 67-DDNQTIENKLEIFVSGYKMNPKIAKKYSVSVKFVDKEKSRAYRLVGVPKAGTGYTLSVW 125   DN+         +G   + KI  KY  SV       +      GV K   G  LS+W PilE 65PKDNTS---------AGVASSDKIKGKYVQSVTVAKGVVTAEMASTGVNKEIQGKKLSLW 115Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF89 shows 83.3% identity over a 162aa overlap with an ORF (ORF89a)from strain A of N. meningitidis:

The complete length ORF89a nucleotide sequence <SEQ ID 341> is:

1 ATGATGAGTA ATAAAATGGA ACAAAAAGGG TTTACATTGA TTGNGANGNT 51 NATNGNCNTCGCGATACNCN GCNTTANCAG CGTCATTNCN ATNNNTNCNT 101 ATCNNAGTTA TATTGAAAAAGGCTATCAGT CCCAGCTTTA TACGGAGATG 151 GTCGGTATCA ACAATATTTC CAAACAGTNTATTTTGAAAA ATCCCCTGGA 201 CGATAATCAG ACCATCAAGA GCAAACTGGA AATATTTGTCTCAGGCTATA 251 AGATGAATCC GAAAATTGCC GAAAAATATA ATGTTTCGGT GCATTTTGTC301 AATGAGGAAA AACCNAGGGC ATACAGCTTG GTCGGCGTTC CAAAGACGGG 351GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC GACGGATACA 401 AATGCCGTGATGCCGCTTCT GCCCGAGCCC ATTTGGAGAC CTTGTCCTCA 451 GATGTCGGCT GTGAAGCCTTCTCTAATCGT AAAAAATAG

This encodes a protein having amino acid sequence <SEQ ID 342>:

1 MMSNKMEQKG FTLIXXXXXX AIXXXXSVIX XXXYXSYIEK GYQSQLYTEM 51 VGINNISKQXILKNPLDDNQ TIKSKLEIFV SGYKMNPKIA EKYNVSVHFV 101 NEEKPRAYSL VGVPKTGTGYTLSVWMNSVG DGYKCRDAAS ARAHLETLSS 151 DVGCEAFSNR KK*

ORF89a and ORF89-1 show 83.3% identity in 162 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF89 shows 84.6% identity over a 162aa overlap with a predicted ORF(ORF89.ng) from N. gonorrhoeae:

The complete length ORF89ng nucleotide sequence <SEQ ID 343> is:

1 aTGATGAGCA ATAAAATGGA ACAAAAAGGG TTTACATTGA TTGAGATGAT 51 GATAGTTGTCACGATACTCG GCATCATCAG CGTCATTGCC ATACCTTCTT 101 ATCAGAGTTA TATTGAAAAAGGCTATCAGT CCCAGCTTTA TACGGAGATG 151 GTCGGTATCA ACAATGTTCT CAAACAGTTTATTTTGAAAA ATCCCCAGGA 201 CGATAATGAT ACCCTCAAGA GCAAACTGAA AATATTTGTCTCAGGCTATA 251 AGATGAATCC GAAAAttgCC AAAAAATATA GTGTTTCGGt aaggtttGTC301 gatGCGGAAA AACCAAGGGC ATACAGGTTG GTCGGCGTTC CGAACGCGGG 351GACGGGTTAT ACTTTGTCGG TATGGATGAA CAGCGTGGGC GACGGATACA 401 AATGCCGTGATGCCACTTCT GCCCAGGCCT ATTCGGACAC CTTGTCCGCA 451 GATAGCGGCT GTGAAGCTTTCTCTAATCGT AAAAAATAG

This encodes a protein having amino acid sequence <SEQ ID 344>:

1 MMSNKMEQ KG FTLIEMMIVV TILGIISVIA IPSYQSYIEK GYQSQLYTEM 51 VGINNVLKQFILKNPQDDND TLKSKLKIFV SGYKMNPKIA KKYSVSVRFV 101 DAEKPRAYRL VGVPNAGTGYTLSVWMNSVG DGYKCRDATS AQAYSDTLSA 151 DSGCEAFSNR KK*

This gonococcal protein has a putative leader peptide (underlined) andN-terminal methylation site (NMePhe or type-4 pili, double-underlined).In addition, ORF89ng and ORF89-1 show 88.3% identity in 162 aa overlap:

Based on this analysis, including the gonococcal motifs and the homologywith the known PilE protein, it was predicted that these proteins fromN. meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF89-1 (13.6 kDa) was cloned in the pGex vector and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 11A shows the results ofaffinity purification of the GST-fusion protein. Purified GST-fusionprotein was used to immunise mice, whose sera gave a positive result inthe ELISA test, confirming that ORF89-1 is a surface-exposed protein,and that it is a useful immunogen.

Example 41

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 345>:

1 ATGAAAAAAT CCTCCCTCAT CAGCGCATTG GGCATCGGTA TTTTGAGCAT 51 CGGCATGGCATTTGCCGCCC CTGCCGACGC GGTAAGCCAA ATCCGTCAAA 101 ACGCCACTCA AGTATTGAGCATCTTAAAAA ACGGCGATGC CAACACCGCT 151 CGCCAAAAAG CCGAAGCCTA TGCGATTCCCTATTTCGATT TCCAACGTAT 201 GACCGCATTG GCGGTCGGCA ACCCTTGGsG CACCG.GTCCGACG.GCAAA 251 AACAAGCGTT GGCCn.AGAA TTTCAACCC...

This corresponds to the amino acid sequence <SEQ ID 346; ORF91>:

1 MKKSSLISAL GIGILSIGMA FAAPADAVSQ IRQNATQVLS ILKNGDANTA 51 RQKAEAYAIPYFDFQRMTAL AVGNPWXTXS DXQKQALAXE FQP...

Further work revealed the complete nucleotide sequence <SEQ ID 347>:

1 ATGAAAAAAT CCTCCCTCAT CAGCGCATTG GGCATCGGTA TTTTGAGCAT 51 CGGCATGGCATTTGCCGCCC CTGCCGACGC GGTAAGCCAA ATCCGTCAAA 101 ACGCCACTCA AGTATTGAGCATCTTAAAAA ACGGCGATGC CAACACCGCT 151 CGCCAAAAAG CCGAAGCCTA TGCGATTCCCTATTTCGATT TCCAACGTAT 201 GACCGCATTG GCGGTCGGCA ACCCTTGGCG CACCGCGTCCGACGCGCAAA 251 AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG CACCTATTCC301 GGCACGATGC TGAAATTAAA AAACGCCAAC GTCAACGTCA AAGACAATCC 351CATCGTCAAT AAAGGCGGCA AAGAAATCAT CGTCCGCGCC GAAGTCGGCG 401 TACCCGGGCAAAAACCCGTC AACATGGACT TCACCACCTA CCAAAGCGGC 451 GGTAAATACC GTACCTACAACGTCGCCATC GAAGGCGCGA GCCTGGTTAC 501 CGTGTACCGC AACCAATTCG GCGAAATTATCAAAGCGAAA GGCGTGGACG 551 GACTGATTGC CGAGTTGAAA GCCAAAAACG GCGGCAAATA A

This corresponds to the amino acid sequence <SEQ ID 348; ORF91-1>:

1 MKKSSLISAL GIGILSIGMA FAAPADAVSQ IRQNATQVLS ILKNGDANTA 51 RQKAEAYAIPYFDFQRMTAL AVGNPWRTAS DAQKQALAKE FQTLLIRTYS 101 GTMLKLKNAN VNVKDNPIVNKGGKEIIVRA EVGVPGQKPV NMDFTTYQSG 151 GKYRTYNVAI EGASLVTVYR NQFGEIIKAKGVDGLIAELK AKNGGK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF91 shows 92.4% identity over a 92aa overlap with an ORF (ORF91a) fromstrain A of N. meningitidis:

The complete length ORF91a nucleotide sequence <SEQ ID 349> is:

1 ATGAAAAAAT CCTCCTTCAT CAGCGCATTG GGCATCGGTA TTTTGAGCAT 51 CGGCATGGCATTTGCCGCCC CTGCCGACGC GGTAAACCAA ATCCGTCAAA 101 ACGCCACTCA AGTATTGAGCATCTTAAAAA GCGGTGATGC CAACACCGCC 151 CGCCAAAAAG CCGAAGCCTA TGCGATTCCCTATTTCGATT TCCAACGTAT 201 GACCGCATTG GCGGTCGGCA ACCCTTGGCG CACCGCGTCCGACGCGCAAA 251 AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG CACCTATTCC301 GGCACGATGC TGAAATTAAA AAACGCCAAC GTCAACGTCA AAGACAATCC 351CATCGTCAAT AAAGGCGGCA AAGAAATCAT CGTCCGCGCC GAAGTCGGCG 401 TACCCGGGCAAAAACCCGTC AACATGGACT TCACCACCTA CCAAAGCGGC 451 GGTAAATACC GTACCTACAACGTCGCCATC GAAGGCGCGA GCCTGGTTAC 501 CGTGTACCGC AACCAATTCG GCGAAATTATCAAAGCGAAA GGCGTGGACG 551 GACTGATTGC CGAGTTGAAG GCTAAAAACG GCAGCAAGTA A

This encodes a protein having amino acid sequence <SEQ ID 350>:

1 MKKSSFISAL GIGILSIGMA FAAPADAVNQ IRQNATQVLS ILKSGDANTA 51 RQKAEAYAIPYFDFQRMTAL AVGNPWRTAS DAQKQALAKE FQTLLIRTYS 101 GTMLKLKNAN VNVKDNPIVNKGGKEIIVRA EVGVPGQKPV NMDFTTYQSG 151 GKYRTYNVAI EGASLVTVYR NQFGEIIKAKGVDGLIAELK AKNGSK*

ORF91a and ORF91-1 show 98.0% identity in 196 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF91 shows 84.8% identity over a 92aa overlap with a predicted ORF(ORF91.ng) from N. gonorrhoeae:

The complete length ORF91ng nucleotide sequence <SEQ ID 351> ispredicted to encode a protein having amino acid sequence <SEQ ID 352>:

1 VKKSSFISAL GIGILSIGMA FASPADAVGQ IRQNATQVLT ILKSGDAASA 51RPKAEAYAVP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE FQTLLIRTYS 101GTMLKFKNAT VNVKDNPIVN KGGKEIVVRA EVGIPGQKPV NMDFTTYQSG 151GKYRTYNVAI EGTSLVTVYR NQFGEIIKAK GIDGLIAELK AKNGGK*

Further work revealed the complete nucleotide sequence <SEQ ID 353>:

1 ATGAAAAAAT CCTCCTTCAT CAGCGCATTG GGCATCGGTA TTTTGAGCAT 51CGGCATGGCA TTTGCCTCCC CGGCCGACGC AGTGGGACAA ATCCGCCAAA 101ACGCCACACA GGTTTTGACC ATCCTCAAAA GCGGCGACGC GGCTTCTGCA 151CGCCCAAAAG CCGAAGCCTA TGCGGTTCCC TATTTCGATT TCCAACGTAT 201GACCGCATTG GCGGTCGGCA ACCCTTGGCG TACCGCGTCC GACGCGCAAA 251AACAAGCGTT GGCCAAAGAA TTTCAAACCC TGCTGATCCG CACCTATTCC 301GGCACGATGC TGAAATTCAA AAACGCGACC GTCAACGTCA AAGACAATCC 351CATCGTCAAT AAGGGCGGCA AGGAAATCGT CGTCCGTGCC GAAGTCGGCA 401TCCCCGGTCA GAAGCCCGTC AATATGGACT TTACCACCTA CCAAAGCGGC 451GGCAAATACC GTACCTACAA CGTCGCCATC GAAGGCACGA GCCTGGTTAC 501CGTGTACCGC AACCAATTCG GCGAAATCAT CAAAGCCAAA GGCATCGACG 551GGCTGATTGC CGAGTTGAAA GCCAAAAACG GCGGCAAATA A

This corresponds to the amino acid sequence <SEQ ID 354; ORF91ng-1>:

1 MKKSSFISAL GIGILSIGMA FASPADAVGQ IRQNATQVLT ILKSGDAASA 51RPKAEAYAVP YFDFQRMTAL AVGNPWRTAS DAQKQALAKE FQTLLIRTYS 101GTMLKFKNAT VNVKDNPIVN KGGKEIVVRA EVGIPGQKPV NMDFTTYQSG 151GKYRTYNVAI EGTSLVTVYR NQFGEIIKAK GIDGLIAELK AKNGGK*

ORF91ng-1 and ORF91-1 show 92.3% identity in 196 aa overlap:

In addition, ORF91ng-1 shows homology to a hypothetical E. coli protein:

sp|P45390|YRBC_ECOLI HYPOTHETICAL 24.0 KD PROTEIN IN MURA-RPONINTERGENICREGION PRECURSOR (F211) >gi|606130 (U18997) ORF_f211 [Escherichia coli] >gi|1789583 (AE000399) hypothetical 24.0 kD protein in murZ-rpoNintergenic region [Escherichia coli]Length = 211 Score =70.6 bits (170), Expect = 6e−12 Identities = 42/137 (30%), Positives =76/137 (54%), Gaps = 6/137 (4%) Query: 59VPYFDFQRMTALAVGNPWRTASDAQKQALAKEFQTLLIRTYSGTMLKFKNATVNVKDNPI 118 +PY   +  AL +G  +++A+ AQ++A    F+  L + Y   +  +   T  +   P Sbjct: 65LPYVQVKYAGALVLGQYYKSATPAQREAYFAAFREYLKQAYGQALAMYHGQTYQIA--PE 122 Query:119 VNKGGKEIV-VRAEVGIP-GQKPVNMDFTTYQSG--GKYRTYNVAIEGTSLVTVYRNQFG 174   G K IV +R  +  P G+ PV +DF   ++   G ++ Y++  EG S++T  +N++G Sbjct: 123QPLGDKTIVPIRVTIIDPNGRPPVRLDFQWRKNSQTGNWQAYDMIAEGVSMITTKQNEWG 182 Query:175 EIIKAKGIDGLIAELKA 191  +++ KGIDGL A+LK+ Sbjct: 183 TLLRTKGIDGLTAQLKS199

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it is predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

Example 42

The following DNA sequence was identified in N. meningitidis <SEQ ID355>:

1 ATGAAACACA TACTCCCCCT GATTGCCGCA TCCGCACTCT GCATTTCAAC 51CGCTTCGGCA CATCCTGCCA GCGAACCGTC CACTCAAAAC GAAACCGCTA 101TGATCACGCA TACCCTCATC TCAAAATACA GTTTTGGnnn nnnnnnnnnn 151nnnnnnnnnn nnGCCATAAA AAGCAAAGGG ATGGACATTT TTGCCGTCAT 201CGACCATCAG GAAGCCGCAC GCCGAAACGG CTTAACGATG CAGCCGGCAA 251AAGTCATCGT CTTCGGCACG CCCAAAGCCG GCACGCCGCT GATGGTCAAA 301GACCCCGCCT TCGCCCTGCA ACTGCCCCTA CGCGTCCTCG TTACCGAAAC 351GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC CTCATCGCCG 401GCAGCCGCAT CGGTTTCGAC GAAGTGGCAA ACACTTTGGC AAACGCCGAA 451AAACTGATAC AAAAAACCGT AGGCGAATAA

This corresponds to the amino acid sequence <SEQ ID 356; ORF97>:

1 MKHILPLIAA SALCISTASA HPASEPSTQN ETAMITHTLI SKYSFGXXXX 51XXXXAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT PKAGTPLMVK 101DPAFALQLPL RVLVTETDGK VRAAYTDTRA LIAGSRIGFD EVANTLANAE 151 KLIQKTVGE*

Further work revealed the complete nucleotide sequence <SEQ ID 357>:

1 ATGAAACACA TACTCCCCCT GATTGCCGCA TCCGCACTCT GCATTTCAAC 51 CGCTTCGGCACATCCTGCCA GCGAACCGTC CACCCAAAAC GAAACCGCTA 101 TGACCACGCA TACCCTCACCTCAAAATACA GTTTTGACGA AACCGTCAGC 151 CGCCTTGAAA CCGCCATAAA AAGCAAAGGGATGGACATTT TTGCCGTCAT 201 CGACCATCAG GAAGCCGCCC GCCGAAACGG CTTAACGATGCAGCCGGCAA 251 AAGTCATCGT CTTCGGCACG CCCAAAGCCG GCACGCCGCT GATGGTCAAA301 GACCCCGCCT TCGCCCTGCA ACTGCCCCTA CGCGTCCTCG TTACCGAAAC 351GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC CTCATCGCCG 401 GCAGCCGCATCGGTTTCGAC GAAGTGGCAA ACACTTTGGC AAACGCCGAA 451 AAACTGATAC AAAAAACCGTAGGCGAATAA

This corresponds to the amino acid sequence <SEQ ID 358; ORF97-1>:

1 MKHILPLIAA SALCISTASA HPASEPSTQN ETAMTTHTLT SKYSFDETVS 51RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT PKAGTPLMVK 101DPAFALQLPL RVLVTETDGK VRAAYTDTRA LIAGSRIGFD EVANTLANAE 151 KLIQKTVGE*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF97 shows 88.7% identity over a 159aa overlap with an ORF (ORF97a)from strain A of N. meningitidis:

The complete length ORF97a nucleotide sequence <SEQ ID 359> is:

1 ATGANACACA TACTCCCCCT GANTGNCGCA TCCGCACTCT GCATTTCAAC 51CGCTTCGGNN CATCCTGCCA GCGAACCGCA AACCCAAAAC GAAACCGCTA 101TGACCACGCA TACCCTCACC TCAAAATACA GTTTTGACGA AACCGTCAGC 151CGCCTTGAAA CCGCCATAAA AAGCAAAGGG ATGGACATTT TTGCCGTCAT 201CGACCATCAG GAAGCCGCCC GCCGAAACGG CTTAACGATG CAGCCGGCAA 251AAGTCATCGT CTTCGGCACG CCCAAAGCCG GTACGCCGCT GATGGTCAAA 301GACCCCGCCT TCGCCCTGCA ACTGCCCCTG CGCGTCNTCG TTACCGAAAC 351GGACGGCAAA GTACGCGCCG CCTATACCGA TACGCGCGCC CTCATCGCCG 401GCAGCCGCAT CGGTTTCGAC GAAGTGGCAA ACACTTTGGC AAACGCCGAA 451AAACTGATAC AAAAAACCAT AGGCGAATAA

This encodes a protein having amino acid sequence <SEQ ID 360>:

1 MXHILPLXXA SALCISTASX HPASEPQTQN ETAMTTHTLT SKYSFDETVS 51RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT PKAGTPLMVK 101DPAFALQLPL RVXVTETDGK VRAAYTDTRA LIAGSRIGFD EVANTLANAE 151 KLIQKTIGE*

ORF97a and ORF97-1 show 95.6% identity in 159 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF97 shows 88.1% identity over a 159aa overlap with a predicted ORF(ORF97.ng) from N. gonorrhoeae:

The complete length ORF97ng nucleotide sequence <SEQ ID 361> ispredicted to encode a protein having amino acid sequence <SEQ ID 362>:

1 MKHILPPIAA SAFCISTASA HPAGKPPTQN ETAMTTHTLT SKYSFDETVS 51RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT PKAGTPLMVK 101DPAFALQLPL RVLVTETDGK VRTAYTDTRA LIVGSRISFD EVANTLANAE 151 KLIQKTVGE*

Further work revealed the complete nucleotide sequence <SEQ ID 363>:

1 ATGAAACACA TACTCCCcct gatcgccgca TccgcactCT GCATTTCAAC 51CGCTTCGGCA CACCCTGCCG GCAAACCGCC CACCCAAAAC GAAACCGCTA 101TGACCACGCA CACCCTCACC TCGAAATACA GTTTTGACGA AACCGTCAGC 151CGCCTTGAAA CCGCCATAAA AAGCAAAGGG ATGGACATTT TTGCCGTCAT 201CGACCATCAG GAAGCGGCAC GCCGAAACGG CCTGACCATG CAGCCGGCAA 251AAGTCATCGT CTTCGGCACG CCCAAGGCCG GTACGCCgct GATGGTCAAA 301GACCCCGCCT TCGCCCTGCA ACTGCCCCTG CGCGTCCTCG TTACCGAAAC 351GGACGGCAAA GTACGCACCG CCTATACCGA TACGCGCGCC CTCATCGTCG 401GCAGCCGCAT CAGTTTCGAC GAAGTGGCAA ACACTTTGGC AAACGCCGAA 451AAACTGATAC AAAAAACCGT AGGCGAATAA

This corresponds to the amino acid sequence <SEQ ID 364; ORF97ng-1>:

1 MKHILPLIAA SALCISTASA HPAGKPPTQN ETAMTTHTLT SKYSFDETVS 51RLETAIKSKG MDIFAVIDHQ EAARRNGLTM QPAKVIVFGT PKAGTPLMVK 101DPAFALQLPL RVLVTETDGK VRTAYTDTRA LIVGSRISFD EVANTLANAE 151 KLIQKTVGE*

ORF97ng-1 and ORF97-1 show 96.2% identity in 159 aa overlap:

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it was predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

ORF97-1 (15.3 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIGS. 12A & 12B show,respectively, the results of affinity purification of the GST-fusion andHis-fusion proteins. Purified GST-fusion protein was used to immunisemice, whose sera were used for Western Blot (FIG. 12C), ELISA (positiveresult), and FACS analysis (FIG. 12D). These experiments confirm thatORF97-1 is a surface-exposed protein, and that it is a useful immunogen.FIG. 12E shows plots of hydrophilicity, antigenic index, and AMPHIregions for ORF97-1.

Example 43

The following DNA, believed to be complete, sequence was identified inN. meningitidis <SEQ ID 365>:

1 ATGGCTTTTA TTACGCGCTT ATTCAAAAGC AGTAAATGGC TGATTGTGCC 51GCTGATGCTC CCCGCCTTTC AGAATGTGGC GGCGGAGGGG ATAGATGTGA 101GCCGTGCCGA AGCGAGGATA ACCGACGGCG GGCAGCTTTC CATCAGCAGC 151CGCTTCCAAA CCGAGCTGCC CGACCAGCTC CAACAGGCGT TGCGCCGGGg 201CGTGCCGCTC AACTTTACCT TAAGCTGGCA GCTTTCCGCC CCGATAATCG 251CTTCTTATCG GTTTAAATTG GGGCAACTGA TTGGCGATGA CGACaATATT 301GACTACAAAC TGAGTTTCCA TCCGCTGACc AaACGCTACC GCGTTACCgT 351CGgCGCGTTT TCGACAGACT ACGACACCTT GGATGCGGCA TTGCGCGCGA 401CCGGCGCGGT TGCCAACTGG AAAGTCCTGA ACAAAGGCGC GCTGTCCGGT 451GCGGAAGCAG GGGAAACCAA GGCGGAAATC CGCCTGACGC TGTCCACTTC 501AAAACTGCCC AAGCCTTTTC AAATCAATGC ATTGACTTCT CAAAACTGGC 551ATTTGGATTC GGGTTGGAAA CCTCTAAACA TCATCGGGAA CAAATAA

This corresponds to the amino acid sequence <SEQ ID 366; ORF106>:

1 MAFITRLFKS SKWLIVPLML PAFQNVAAEG IDVSRAEARI TDGGQLSISS 51RFQTELPDQL QQALRRGVPL NFTLSWQLSA PIIASYRFKL GQLIGDDDNI 101DYKLSFHPLT KRYRVTVGAF STDYDTLDAA LRATGAVANW KVLNKGALSG 151AEAGETKAEI RLTLSTSKLP KPFQINALTS QNWHLDSGWK PLNIIGNK*

Further work revealed the following DNA sequence <SEQ ID 367>:

1 ATGGCTTTTA TTACGCGCTT ATTCAAAAGC AGTAAATGGC TGATTGTGCC 51 GCTGATGCTCCCCGCCTTTC AGAATGTGGC GGCGGAGGGG ATAGATGTGA 101 GCCGTGCCGA AGCGAGGATAACCGACGGCG GGCAGCTTTC CATCAGCAGC 151 CGCTTCCAAA CCGAGCTGCC CGACCAGCTCCAACAGGCGT TGCGCCGGGG 201 CGTGCCGCTC AACTTTACCT TAAGCTGGCA GCTTTCCGCCCCGATAATCG 251 CTTCTTATCG GTTTAAATTG GGGCAACTGA TTGGCGATGA CGACAATATT301 GACTACAAAC TGAGTTTCCA TCCGCTGACC AACCGCTACC GCGTTACCGT 351CGGCGCGTTT TCGACAGACT ACGACACCTT GGATGCGGCA TTGCGCGCGA 401 CCGGCGCGGTTGCCAACTGG AAAGTCCTGA ACAAAGGCGC GCTGTCCGGT 451 GCGGAAGCAG GGGAAACCAAGGCGGAAATC CGCCTGACGC TGTCCACTTC 501 AAAACTGCCC AAGCCTTTTC AAATCAATGCATTGACTTCT CAAAACTGGC 551 ATTTGGATTC GGGTTGGAAA CCTCTAAACA TCATCGGGAACAAATAA

This corresponds to the amino acid sequence <SEQ ID 368; ORF106-1>:

1 MAFITRLFKS SKWLIVPLML PAFQNVAAEG IDVSRAEARI TDGGQLSISS 51 RFQTELPDQLQQALRRGVPL NFTLSWQLSA PIIASYRFKL GQLIGDDDNI 101 DYKLSFHPLT NRYRVTVGAFSTDYDTLDAA LRATGAVANW KVLNKGALSG 151 AEAGETKAEI RLTLSTSKLP KPFQINALTSQNWHLDSGWK PLNIIGNK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF106 shows 87.4% identity over a 199aa overlap with an ORF (ORF106a)from strain A of N. meningitidis:

Due to the K→N substitution at residue 111, the homology between ORF106aand ORF106-1 is 87.9% over the same 199 aa overlap.

The complete length ORF106a nucleotide sequence <SEQ ID 369> is:

1 ATGGCTTTTA TTACGCGCTT ATTCAAAAGC ATTAAACAAT GGCTTGTGCT 51 GCTGCCGATGCTTTCCGTTT TGCCGGACGC GGCGGCGGAG GGGATAGATG 101 TGAGCCGCGC CGAAGCGAGGATAANCGACG GCGGGCAGCT TTCCATNAGN 151 AGCCGCTTCC AAACCGAGCT GCCCGACCAGCTCCAANNNG CGNNGNGCCG 201 GGGCGTGNCG CTCAACTNTA CCTTAAGNTG GCAGCTTTCCGCCCCGATAA 251 TCGCTTCTTA TCGGTTTNAA TTGGGGCAAC TGATTGGCGA TGACGACNAT301 ATTGACTACA AACTGAGTTT CCATCCGCTG ACCAACCGCT ACCGCGTTAC 351CGTCGGCGCG TTTTCGACAG ANTACGACAC CTTGGATGCG GCATTGCGCG 401 CGACCGGCGCGGTTGCCAAC TGGAAAGTCC TGAACAAAGG CGCGCTGTCC 451 GGTGCGGAAG CAGGGGAAACCAAGGCGGAA ATCCGCCTGA CGCTGTCCAC 501 TTCAAAACTG CCCAAGCCTT TTCAAATCAATGCATTGACT TCTCAAAACT 551 GGCATTTGGA TTCGGGTTGG AAACCTCTAA ACATCATCGGGAACAAATAA

This encodes a protein having amino acid sequence <SEQ ID 370>:

1 MAFITRLFKS IKQWLVLLPM LSVLPDAAAE GIDVSRAEAR IXDGGQLSXX 51 SRFQTELPDQLQXAXXRGVX LNXTLXWQLS APIIASYRFX LGQLIGDDDX 101 IDYKLSFHPL TNRYRVTVGAFSTXYDTLDA ALRATGAVAN WKVLNKGALS 151 GAEAGETKAE IRLTLSTSKL PKPFQINALTSQNWHLDSGW KPLNIIGNK*Homology with a Predicted ORF from N. gonorrhoeae

ORF106 shows 90.5% identity over a 199aa overlap with a predicted ORF(ORF106.ng) from N. gonorrhoeae:

Due to the K→N substitution at residue 111, the homology betweenORF106ng and ORF106-1 is 91.0% over the same 199 aa overlap.

The complete length ORF106ng nucleotide sequence <SEQ ID 371> is:

1 ATGGCTTTTA TTACGCGCTT ATTCAAAAGC ATTAAACAAT GGCTTGTGCT 51 GTTGCCGATACTCTCCGTTT TGCCGGACGC GGCGGCGGAG GGCATTGCCG 101 CGACCCGCGC CGAAGCGAGGATAACCGACG GCGGGCGGCT TTCCATCAGC 151 AGCCGCTTCC AAACCGAGCT GCCCGACCAGCTCCAACAGG CGTTGCGCCG 201 GGGCGTACCG CTCAACTTTA CCTTAAGCTG GCAGCTTTCCGCCCCGACAA 251 TCGCTTCTTA TCGGTTTAAA TTGGGGCAAC TGATTGGCGA TGACGACAAT301 ATTGACTACA AACTAAGTTT CCATCCGCTG ACCAACCGCT ACCGCGTTAC 351CGTCGGCGCA TTTTCCACCG ATTACGACAC TTTGGATGCG GCATTGCGCG 401 CGACCGGCGCGGTTGCCAAC TGGAAAGTCC TGAACAAAGG CGCGTTGTCC 451 GGTGCGGAAG CAGGGGAAACCAAGGCGGAA ATCCGCCTGA CGCTGTCCAC 501 TTCAAAACTG CCCAAGCCTT TCCAAATCAACGCATTGACT TCTCAAAACT 551 GGCATTTGGA TTCGGGTTGG AAACCTCTAA ACATCATCGGGAACAAATAA

This encodes a protein having amino acid sequence <SEQ ID 372>:

1 MAFITRLFKS IKQWLVLLPI LSVLPDAAAE GIAATRAEAR ITDGGRLSIS 51 SRFQTELPDQLQQALRRGVP LNFTLSWQLS APTIASYRFK LGQLIGDDDN 101 IDYKLSFHPL TNRYRVTVGAFSTDYDTLDA ALRATGAVAN WKVLNKGALS 151 GAEAGETKAE IRLTLSTSKL PKPFQINALTSQNWHLDSGW KPLNIIGNK*

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it was predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

ORF106-1 (18 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 13A shows the results ofaffinity purification of the His-fusion protein, and FIG. 13B shows theresults of expression of the GST-fusion in E. coli. Purified His-fusionprotein was used to immunise mice, whose sera were used for FACSanalysis (FIG. 13C) These experiments confirm that ORF106-1 is asurface-exposed protein, and that it is a useful immunogen.

Example 44

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 373>:

1 ATGGACACAA AAGAAATCCT CGG.TACGCG GcAGGcTCGA TCGGCAGCGC 51 GGTTTTAGCCGTCATCATCc TGCCGCTGCT GTCGTGGTAT TTCCCCGCCG 101 ACGACATCGG GCGCATCGTGCTGATGCAGA CGGCGGCGGG GCTgACGGTG 151 TCGGTGTTGT GCCTCGGGCT GGATCAGGCATACGTCCGCG AATACTATGC 201 CACCGCCGAC AAAGACAcCT TGTTCAAAAC CCTGTTCCTGCCGCCGCTGC 251 TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC GTCCCTGCCG301 TCTGAAATCC TGTTTTCACT CGACGATGCC gCCGCCGGCa TCGGGCTGGT 351GCTGTTTGAA CtGAGCTTCC TGCCCATCCG cTTTCTCTTA CTGGTTTTGC 401 GTATGGAAGGACGCGCCcTT GCCTTTTCGT CCGCGCAACT CGTGCcCAAG 451 CTCGCCATCC TGCTGCTG.TGCCGCTGACG GTCGGGCTGC TGCACTTTCC 501 AGCGAACACC GCCGTCCTGA CCGCCGTTTACGCGCTGGCA AACCTTGCCG 551 CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAAGGCCGTCCGG 601 CACGCACCGT TTTCGCCCGC CGTCCTGCAC CGGGGG.TGC GCTACGGCAT651 ACCGATCGCA CTGAGCAGCA TCGCCTATTG GGGGCTGGCA TCCGCCGACC 701GTTTGTTCCT GAAAAAATAT GCCGGCCTGG AACAGCTCGG CGTTTATTCG 751 ATGGGTATTTCGTTCGGCGG GGCGGCATTA TTGTTCCAAA GCATCTTTTC 801 AACGGTCTGG ACACCGTATATTTTCCGCGC AATCGAAGAA AACGCCCCGC 851 CCGCTCGCCT CTCGGCAACG GCAGAATCCGCCGCCGCCCT GCTTGCCTCC 901 GCCCTCTGC. TGACCGGCAT TTTCTCGCCC CTTGCCTCCCTCCTGCTGCC 951 GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT ATG.TGCCGC1001 CGCTGTTTTG CACGCTGGCG GAAATCAGCG GCATCGGTTT GAACGTCGTT 1051CGCAAAACGC GCCCGATCGC GCTCGCCACC TTGGGCGCGC TGGCGGCAAA 1101 CCTGCTGCTGCTGGGGCTTG ACCGTGCCGT ACCGGCGAGG CCGCC.GGCG 1151 CGGCGGTTGC CTGTGCCGCCTCATTCTGGC TGTTTTTTGC CTTCAAGACC 1201 GAAAGCTCyT GCCGCCTGTG GCAGCCGCTCAAACGCCTGC CGCTTTATCT 1251 GCACACATTG TTCTGCCTGA CCTCCTCGGC GGCCTACACCTGCTTCGGCA 1301 CGCCGGCAAA CTATCCCCTG TTTGCCGGCG TATGGGCGGC ATATCTGGCA1351 GGCTGCATCC TGCGCCACCG GAAAGATTTG CACAAACTGT TTCATTATTT 1401GAAAAAACAA GGTTTCCCAT TATGA

This corresponds to the amino acid sequence <SEQ ID 374; ORF10>:

1 MDTKEILXYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV LMQTAAGLTV 51 SVLCLGLDQAYVREYYATAD KDTLFKTLFL PPLLSAAAIA ALLLSRPSLP 101 SEILFSLDDA AAGIGLVLFELSFLPIRFLL LVLRMEGRAL AFSSAQLVPK 151 LAILLLXPLT VGLLHFPANT AVLTAVYALANLAAAAFLLF QNRCRLKAVR 201 HAPFSPAVLH RGXRYGIPIA LSSIAYWGLA SADRLFLKKYAGLEQLGVYS 251 MGISFGGAAL LFQSIFSTVW TPYIFRAIEE NAPPARLSAT AESAAALLAS301 ALCXTGIFSP LASLLLPENY AAVRFIVVSC MXPPLFCTLA EISGIGLNVV 351RKTRPIALAT LGALAANLLL LGLDRAVPAR PXGAAVACAA SFWLFFAFKT 401 ESSCRLWQPLKRLPLYLHTL FCLTSSAAYT CFGTPANYPL FAGVWAAYLA 451 GCILRHRKDL HKLFHYLKKQGFPL*

Further sequence analysis revealed the complete DNA sequence <SEQ ID375> to be:

1 ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA TCGGCAGCGC 51 GGTTTTAGCCGTCATCATCC TGCCGCTGCT GTCGTGGTAT TTCCCCGCCG 101 ACGACATCGG GCGCATCGTGCTGATGCAGA CGGCGGCGGG GCTGACGGTG 151 TCGGTGTTGT GCCTCGGGCT GGATCAGGCATACGTCCGCG AATACTATGC 201 CACCGCCGAC AAAGACACCT TGTTCAAAAC CCTGTTCCTGCCGCCGCTGC 251 TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC GTCCCTGCCG301 TCTGAAATCC TGTTTTCACT CGACGATGCC GCCGCCGGCA TCGGGCTGGT 351GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA CTGGTTTTGC 401 GTATGGAAGGACGCGCCCTT GCCTTTTCGT CCGCGCAACT CGTGCCCAAG 451 CTCGCCATCC TGCTGCTGCTGCCGCTGACG GTCGGGCTGC TGCACTTTCC 501 AGCGAACACC GCCGTCCTGA CCGCCGTTTACGCGCTGGCA AACCTTGCCG 551 CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAAGGCCGTCCGG 601 CACGCACCGT TTTCGCCCGC CGTCCTGCAC CGGGGGCTGC GCTACGGCAT651 ACCGATCGCA CTGAGCAGCA TCGCCTATTG GGGGCTGGCA TCCGCCGACC 701GTTTGTTCCT GAAAAAATAT GCCGGCCTGG AACAGCTCGG CGTTTATTCG 751 ATGGGTATTTCGTTCGGCGG GGCGGCATTA TTGTTCCAAA GCATCTTTTC 801 AACGGTCTGG ACACCGTATATTTTCCGCGC AATCGAAGAA AACGCCCCGC 851 CCGCCCGCCT CTCGGCAACG GCAGAATCCGCCGCCGCCCT GCTTGCCTCC 901 GCCCTCTGCC TGACCGGCAT TTTCTCGCCC CTTGCCTCCCTCCTGCTGCC 951 GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT ATGCTGCCGC1001 CGCTGTTTTG CACGCTGGCG GAAATCAGCG GCATCGGTTT GAACGTCGTC 1051CGCAAAACGC GCCCGATCGC GCTCGCCACC TTGGGCGCGC TGGCGGCAAA 1101 CCTGCTGCTGCTGGGGCTTG CCGTGCCGTC CGGCGGCGCG CGCGGCGCGG 1151 CGGTTGCCTG TGCCGCCTCATTCTGGCTGT TTTTTGCCTT CAAGACCGAA 1201 AGCTCCTGCC GCCTGTGGCA GCCGCTCAAACGCCTGCCGC TTTATCTGCA 1251 CACATTGTTC TGCCTGACCT CCTCGGCGGC CTACACCTGCTTCGGCACGC 1301 CGGCAAACTA TCCCCTGTTT GCCGGCGTAT GGGCGGCATA TCTGGCAGGC1351 TGCATCCTGC GCCACCGGAA AGATTTGCAC AAACTGTTTC ATTATTTGAA 1401AAAACAAGGT TTCCCATTAT GA

This corresponds to the amino acid sequence <SEQ ID 376; ORF10-1>:

1 MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV LMQTAAGLTV 51 SVLCLGLDQAYVREYYATAD KDTLFKTLFL PPLLSAAAIA ALLLSRPSLP 101 SEILFSLDDAAAGIGLVLFE LSFLPIRFLL LVLRMEGRAL AFSSAQLVPK 151 LAILLLLPLT VGLLHFPANTAVLTAVYALA NLAAAAFLLF QNRCRLKAVR 201 HAPFSPAVLH RGLRYGIPIA LSSIAYWGLASADRLFLKKY AGLEQLGVYS 251 MGISFGGAAL LFQSIFSTVW TPYIFRAIEE NAPPARLSATAESAAALLAS 301 ALCLTGIFSP LASLLLPENY AAVRFIVVSC MLPPLFCTLA EISGIGLNVV351 RKTRPIALAT LGALAANLLL LGLAVPSGGA RGAAVACAAS FWLFFAFKTE 401SSCRLWQPLK RLPLYLHTLF CLTSSAAYTC FGTPANYPLF AGVWAAYLAG 451 CILRHRKDLHKLFHYLKKQG FPL*

Computer analysis of this amino acid sequence gave the followingresults:

Prediction

ORF10-1 is predicted to be the precursor of an integral membraneprotein, since it comprises several (12-13) potential transmembranesegments, and a probable cleavable signal peptide

Homology with EpsM from Streptococcus thermophilus (Accession NumberU40830).

ORF10 shows homology with the epsM gene of S. thermophilus, whichencodes a protein of a size similar to ORF10 and is involved inexpolysaccharide synthesis. Other homologies are with prokaryoticmembrane proteins:

Identities = (25%) Query: 213LRYGIPLALSSLAYWGLASADRLFLKKYAGLEQLGVYSMGISFGGAALLLQSIFSTVW 270 L Y+PL  SS+ +W L ++ R F+  + G    G+ ++         +  +IF+  W Sbjct: 210LYYALPLIPSSILWWLLNASSRYFVLFFLGAGANGLLAVATKIPSIISIFNTIFTQAW 267Identities = 15/57 (26%), Positives = 31/57 (54%) Query: 7LGYAAGSIGSAVLAVIILPLLSWYFPADDIGRIVLMQTAAGLTVSVLCLGLDQAYVR 63 L +  G++GS+L  +++PL ++     + G   L QT A L + ++ + +  A +R Sbjct: 12LVFTIGNLGSKLLVFLLVPLYTYAMTPQEYGMADLYQTTANLLLPLITMNVFDATLR 68 Identities= 16/96 (16%), Positives = 36/96 (37%) Query: 307IFSPLASLLLPENYAAVRFTVVSCMLPPLFYTLTEISGIGLNVVRKTRPIXXXXXXXXXX 366  +  P+   ++  +YA+    V   ML  LF + ++  G      ++T+ + Sbjct: 305VLKPIVEKVVSSDYASSWQYVPFFMLSMLFSSFSDFFGTNYIAAKQTKGVFMTSIYGTIV 364Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF10 shows 95.4% identity over a 475aa overlap with an ORF (ORF10a)from strain A of N. meningitidis:

The complete length ORF10a nucleotide sequence <SEQ ID 377> is:

1 ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA TCGGCAGCGC 51 GGTTTTAGCCGTCATCATCC TGCCGCTGCT GTCGTGGTAT TTCCCTGCCG 101 ACGACATCGG ACGCATCGTGCTGATGCAGA CGGCGGCGGG GCTGACGGTG 151 TCGGTGTTGT GCCTCGGGCT GGATCAGGCATACGTCCGCG AATACTATGC 201 CGCCGCCGAC AAAGACACTT TGTTCAAAAC CCTGTTCCTGCCGCCGCTGC 251 TGTCTGCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC ATCCCTGCCG301 TCTGAAATCC TGTTTTCGCT CGACGATGCC GCCGCCGGCA TCGGGCTGGT 351GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA CTGGTTTTGC 401 GTATGGAAGGACGCGCCCTT GCCTTTTCGT CCGCGCAACT CGTGTCCAAG 451 CTCGCCATCC TGCTGCTGCTGCCGCTGACG GTCGGGCTGC TGCACTTTCC 501 GGCGAACACC GCCGTCCTGA CCGCCGTTTACGCGCTGGCA AACCTTGCCG 551 CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAAGGCCGTCCGG 601 CGCGCACCGT TTTCATCCGC CGTCCTGCAT CGCGGCCTGC GCTACGGCAT651 ACCGATCGCA CTAAGCAGCA TCGCCTATTG GGGGCTGGCA TCCGCCGACC 701GTTTGTTCCT GAAAAAATAT GCCGGCCTAG AACAGCTCGG CGTTTATTCG 751 ATGGGTATTTCGTTCGGCGG AGCGGCATTA TTGTTCCAAA GCATCTTTTC 801 AACGGTCTGG ACACCGTATATTTTCCGCGC AATCGAAGCA AACGCCCCGC 851 CCGCCCGCCT CTCGGCAACG GCAGAATCCGCCGCCGCCCT GCTTGCCTCC 901 GCCCTCTGCC TGACCGGCAT TTTCTCGCCC CTCGCCTCCCTCCTGCTGCC 951 GGAAAACTAC GCCGCCGTCC GGTTTATCGT CGTATCGTGT ATGCTGCCTC1001 CGCTGTTTTG CACGCTGGTA GAAATCAGCG GCATCGGTTT GAACGTCGTC 1051CGAAAAACAC GCCCGATCGC GCTCGCCACC TTGGGCGCGC TGGCGGCAAA 1101 CCTGCTGCTGCTGGGGCTTG CCGTACCGTC CGGCGGCGCG CGCGGCGCGG 1151 CGGTTGCCTG TGCCGCCTCATTTTGGCTGT TTTTTGTTTT CAAGACCGAA 1201 AGCTCCTGCC GCCTGTGGCA GCCGCTCAAACGCCTGCCGC TTTATATGCA 1251 CACATTGTTC TGCCTGGCCT CCTCGGCGGC CTACACCTGCTTCGGCACTC 1301 CGGCAAACTA CCCCCTGTTT GCCGGCGTAT GGGCGGTATA TCTGGCAGGC1351 TGCATCCTGC GCCACCGGAA AGATTTGCAC AAACTGTTTC ATTATTTGAA 1401AAAACAAGGT TTCCCATTAT GA

This encodes a protein having amino acid sequence <SEQ ID 378>:

1 MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV LMQTAAGLTV 51 SVLCLGLDQAYVREYYAAAD KDTLFKTLFL PPLLSAAAIA ALLLSRPSLP 101 SEILFSLDDA AAGIGLVLFELSFLPIRFLL LVLRMEGRAL AFSSAQLVSK 151 LAILLLLPLT VGLLHFPANT AVLTAVYALANLAAAAFLLF QNRCRLKAVR 201 RAPFSSAVLH RGLRYGIPIA LSSIAYWGLA SADRLFLKKYAGLEQLGVYS 251 MGISFGGAAL LFQSIFSTVW TPYIFRAIEA NAPPARLSAT AESAAALLAS301 ALCLTGIFSP LASLLLPENY AAVRFIVVSC MLPPLFCTLV EISGIGLNVV 351RKTRPIALAT LGALAANLLL LGLAVPSGGA RGAAVACAAS FWLFFVFKTE 401 SSCRLWQPLKRLPLYMHTLF CLASSAAYTC FGTPANYPLF AGVWAVYLAG 451 CILRHRKDLH KLFHYLKKQGFPL*

ORF10a and ORF10-1 show 95.4% identity in 475 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF10 shows 94.1% identity over a 475aa overlap with a predicted ORF(ORF10.ng) from N. gonorrhoeae:

The complete length ORF10ng nucleotide sequence <SEQ ID 379> is:

1 ATGGACACAA AAGAAATCCT CGGCTACGCG GCAGGCTCGA TCGGCAGCGC 51 GGTTTTAGCCGTCATCATCC TGCCGCTGCT GTCGTGGTAT TTCcccgCCG 101 ACGACATCGG GCGCATCGTGCTGATGCAGA CGGCGGCGGG ACTGACGGTG 151 TCGGTATTGT GCCTCGGGCT GGATCAGGCATACGTCCGCG AATACTATGC 201 CGCCGCCGAC AAAGACACTT TGTTCAAAAC CCTGTTCCTGCCGCCGCTGC 251 TGTTTTCCGC CGCGATAGCC GCCCTGCTGC TTTCCCGCCC GTCCCTGCCG301 TCTGAAATCC TGTTTTCGCT CGACGATGCC GCCGCCGGCA TCGGGCTGGT 351GCTGTTTGAA CTGAGCTTCC TGCCCATCCG CTTTCTCTTA CTGGTTTTGC 401 GTATGGAAGGGCGCGCCCTT GCCTTTTCGT CCGCGCAACT CGTGCCCAAA 451 CTCGCCATTC TGCTGCTGTTGCCGCTGACG GTCGGGCTGC TGCACTTTCC 501 GGCGAACACC TCCGTCCTGA CCGCCGTTTACGCGCTGGCA AACCTTGCCG 551 CCGCCGCCTT TTTGCTGTTT CAAAACCGAT GCCGTCTGAAGGCCGTCCGG 601 CGCGCGCCGT TTTCGCCCGC CGTCCTGCAC CGGGGGCTGC GCTACGGCAT651 ACCGCTCGCA CTGAGCAGCC TTGCCTATTG GGGGCTGGCA TCCGCCGACC 701GTTTGTTCCT GAAAAAATAT GCGGGCCTGG AACAGCTCGG CGTTTATTCG 751 ATGGGTATTTCGTTCGGCGG GGCGGCATTA TTGCTCCAAA GCATCTTTTC 801 AACGGTCTGG ACACCGTATATTTTCCGTGC AATCGAAGAA AACGCCACGC 851 CCGCCCGCCT CTCGGCAACG GCAGAATCCGCCGCCGCCCT GCTTGCCTCC 901 GCCCTCTGCC TGACCGGAAT TTTCTCGCCC CTCGCCTCCCTCCTGCTGCC 951 GGAAAACTAC GCCGCCGTCC GGTTTACCGT CGTATCGTGT ATGCTGccgc1001 cgctGTTTTA CACGCTGACC GAAATCAGCG GCATCGGTTT GAACGTCGTC 1051CGCAAAACGC GTCCGATCGC GCTTGCCACC TTGGGCGCGC TGGCGGCAAA 1101 CCTGCTGCTGCTGGGGCTTG CCGTACCGTC CGGCGGCACG CGCGGCGCGG 1151 CGGTTGCCTG TGCCGCCTCATTCTGGTTGT TTTTTGTTTT CAAGACAGAA 1201 AGCTCCTGCC GCCTGTGGCA GCCGCTCAAACGCCTGCCGC TTTATATGCA 1251 CACATTGTTC TGCCTgGCCT CCTCGGCGGC CTACACCTGCTTCGGCACAC 1301 CGGCAAACTA CCCcctgttt gccggcgtAT GGGCGGCATA TCTGGCAGGC1351 TGCATCCTGC GCCACCGGAA AAATTTGCAC AAACTGTTTC ATTATTTGAA 1401AAAACAAGGT TTCCCATTAT GA

This encodes a protein having amino acid sequence <SEQ ID 380>:

1 MDTKEILGYA AGSIGSAVLA VIILPLLSWY FPADDIGRIV LMQTAAGLTV 51 SVLCLGLDQAYVREYYAAAD KDTLFKTLFL PPLLFSAAIA ALLLSRPSLP 101 SEILFSLDDAAAGIGLVLFE LSFLPIRFLL LVLRMEGRAL AFSSAQLVPK 151 LAIL LLLPLT VGLLHFPANTSVLTAVYALA NLAAAAFLLF QNRCRLKAVR 201 RAPFSPAVLH RGLRYGIPLA LSSLAYWGLASADRLFLKKY AGLEQLGVYS 251 MGISFGGAAL LLQSIFSTVW TPYIFRAIEE NATPARLSATAESAAALLAS 301 ALCLTGIFSP LASLLLPENY AAVRFTVVSC MLPPLFYTLT EISGIGLNVV351 RKTRPIALAT LGALAANLLL LGLAVPSGGT RGAAVACAAS FWLFFVFKTE 401SSCRLWQPLK RLPLYMHTLF CLASSAAYTC FGTPANYPLF AGVWAAYLAG 451 CILRHRKNLHKLFHYLKKQG FPL*

ORF10ng and ORF10-1 show 96.4% identity in 473 aa overlap:

Based on this analysis, including the presence of a putative leaderpeptide and several transmembrane segments and the presence of aleucine-zipper motif(4 Leu residues spaced by 6 aa, shown in bold), itis predicted that these proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 45

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 381>:

1 ..ATCCTGAAAC CGCATAACCA GCTTAAGGAA GACATCCAAC CTGATCCGGC 51  CGATCAAAAC GCCTTGTCCG AACCGGATGC TGCGACAGAG GCAGAGCAGT 101  CGGATGCGGA AAATGCTGCC GACAAGCAGC CCGTTGCCGA TAAAGCCGAC 151  GAGGTTGAAG AAAAGGCGGG CGAGCCGGAA CGGGAAGAGC CGGACGGACA 201  GGCAGTGCGT AAGAAAGCGC TGACGGAAGA GCGTGAACAA ACCGTCAGGG 251  AAAAAGCGCA GAAGAAAGAT GCCGAAACGG TTAAAATACA AGCGGTAAAA 301  CCGTCTAAAG AAACAGAGAA AAAAGCTTCA AAAGAAGAGA AAAAGGCGGC 351  GAAGGAAAAA GTTGCACCCA AACCAACCCC GGAACAAATC CTCAACAGCG 401  GCAgCATCGA AAAmGCGCGC AgTGCCGCCG CCAAAGAAGT GCAGAAAATG 451  AA.AACGTCC GACAAGGCGG AAGC.AACGC ATTATCTGCA AATGGGCGCG 501  TATGCCGACC GTCAGAGCGC GGAAGGGCAG CGTGCCAAAC TGGCAATCTT 551  GGGCATATCT TCCAAGGTGG TCGGTTATCA GGCGGGACAT AAAACGCTTT 601  ACCGGGTGCA AAGCGGCAAT ATGTCTGCCG ATGCGGTGA

This corresponds to the amino acid sequence <SEQ ID 382; ORF65>:

1 ..ILKPHNQLKE DIQPDPADQN ALSEPDAATE AEQSDAENAA DKQPVADKAD 51  EVEEKAGEPE REEPDGQAVR KKALTEEREQ TVREKAQKKD AETVKIQAVK 101  PSKETEKKAS KEEKKAAKEK VAPKPTPEQI LNSGSIEXAR SAAAKEVQKM 151  XNVRQGGSXR IICKWARMPT VRARKGSVPN WQSWAYLPRW SVIRRDIKRF 201  TGCKAAICLP MR*

Further work revealed the complete nucleotide sequence <SEQ ID 383>:

1 ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT CCGGTTTTTT 51 CTTCGGTTTGATACTGGCGA CGGTCATTAT TGCCGGTATT TTGTTTTATC 101 TGAACCAGAG CGGTCAAAATGCGTTCAAAA TCCCGGCTTC GTCGAAGCAG 151 CCTGCAGAAA CGGAAATCCT GAAACCGAAAAACCAGCCTA AGGAAGACAT 201 CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCGGATGCTGCGA 251 CAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA GCAGCCCGTT301 GCCGATAAAG CCGACGAGGT TGAAGAAAAG GCGGGCGAGC CGGAACGGGA 351AGAGCCGGAC GGACAGGCAG TGCGTAAGAA AGCGCTGACG GAAGAGCGTG 401 AACAAACCGTCAGGGAAAAA GCGCAGAAGA AAGATGCCGA AACGGTTAAA 451 AAACAAGCGG TAAAACCGTCTAAAGAAACA GAGAAAAAAG CTTCAAAAGA 501 AGAGAAAAAG GCGGCGAAGG AAAAAGTTGCACCCAAACCA ACCCCGGAAC 551 AAATCCTCAA CAGCGGCAGC ATCGAAAAAG CGCGCAGTGCCGCCGCCAAA 601 GAAGTGCAGA AAATGAAAAC GTCCGACAAG GCGGAAGCAA CGCATTATCT651 GCAAATGGGC GCGTATGCCG ACCGTCAGAG CGCGGAAGGG CAGCGTGCCA 701AACTGGCAAT CTTGGGCATA TCTTCCAAGG TGGTCGGTTA TCAGGCGGGA 751 CATAAAACGCTTTACCGGGT GCAAAGCGGC AATATGTCTG CCGATGCGGT 801 GAAAAAAATG CAGGACGAGTTGAAAAAACA TGAAGTCGCC AGCCTGATCC 851 GTTCTATCGA AAGCAAATAA

This corresponds to the amino acid sequence <SEQ ID 384; ORF65-1>:

1 MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LFYLNQSGQN AFKIPASSKQ 51 PAETEILKPKNQPKEDIQPE PADQNALSEP DAATEAEQSD AEKAADKQPV 101 ADKADEVEEK AGEPEREEPDGQAVRKKALT EEREQTVREK AQKKDAETVK 151 KQAVKPSKET EKKASKEEKK AAKEKVAPKPTPEQILNSGS IEKARSAAAK 201 EVQKMKTSDK AEATHYLQMG AYADRQSAEG QRAKLAILGISSKVVGYQAG 251 HKTLYRVQSG NMSADAVKKM QDELKKHEVA SLIRSIESK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF65 shows 92.0% identity over a 150aa overlap with an ORF (ORF65a)from strain A of N. meningitidis:

The complete length ORF65a nucleotide sequence <SEQ ID 385> is:

1 ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT CCGGTTTTTT 51 CTTCGGTTTGATACTGGCGA CGGTCATTAT TGCCGGTATT TTGTTTTATC 101 TGAACCAGAG CGGTCAAAATGCGTTCAAAA TCCCGGTTCC GTCGAAGCAG 151 CCTGCAGAAA CGGAAATCCT GAAACCGAAAAACCAGCCTA AGGAAGACAT 201 CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCGGATGCTGCGA 251 AAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA GCAGCCCGTT301 GCCGACAAAG CCGACGAGGT TGAGGAAAAG GCGGACGAGC CGGAGCGGGA 351AAAGTCGGAC GGACAGGCAG TGCGCAAGAA AGCACTGACG GAAGAGCGTG 401 AACAAACCGTCGGGGAAAAA GCGCAGAAGA AAGATGCCGA AACGGTTAAA 451 AAACAAGCGG TAAAACCATCTAAAGAAACA GAGAAAAAAG CTTCAAAAGA 501 AGAGAAAAAG GCGGAGAAGG AAAAAGTTGCACCCAAACCG ACCCCGGAAC 551 AAATCCTCAA CAGCGGCAGC ATCGAAAAAG CGCGCAGTGCCGCTGCCAAA 601 GAAGTGCAGA AAATGAAAAC GCCCGACAAG GCGGAAGCAA CGCATTATCT651 GCAAATGGGC GCGTATGCCG ACCGCCGGAG CGCGGAAGGG CAGCGTGCCA 701AACTGGCAAT CTTGGGCATA TCTTCCAAGG TGGTCGGTTA TCAGGCGGGA 751 CATAAAACGCTTTACCGGGT GCAAAGCGGC AATATGTCTG CCGATGCGGT 801 GAAAAAAATG CAGGACGAGTTGAAAAAACA TGAAGTCGCC AGCCTGATCC 851 GTTCTATCGA AAGCAAATAA

This encodes a protein having amino acid sequence <SEQ ID 386>:

1 MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LFYLNQSGQN AFKIPVPSKQ 51 PAETEILKPKNQPKEDIQPE PADQNALSEP DAAKEAEQSD AEKAADKQPV 101 ADKADEVEEK ADEPEREKSDGQAVRKKALT EEREQTVGEK AQKKDAETVK 151 KQAVKPSKET EKKASKEEKK AEKEKVAPKPTPEQILNSGS IEKARSAAAK 201 EVQKMKTPDK AEATHYLQMG AYADRRSAEG QRAKLAILGISSKVVGYQAG 251 HKTLYRVQSG NMSADAVKKM QDELKKHEVA SLIRSIESK*

ORF65a and ORF65-1 show 96.5% identity in 289 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF65 shows 89.6% identity over a 212aa overlap with a predicted ORF(ORF65.ng) from N. gonorrhoeae:

An ORF65ng nucleotide sequence <SEQ ID 387> was predicted to encode aprotein having amino acid sequence <SEQ ID 388>:

1 MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LLYLNQGGQN AFKIPAPSKQ 51 PAETEILKLKNQPKEDIQPE PADQNALSEP DVAKEAEQSD AEKAADKQPV 101 ADKADEVEEK AGEPEREEPDGQAVRKKALT EEREQTVREK AQKKDAETVK 151 KKAVKPSKET EKKASKEEKK AAKEKVAPKPTPEQILNSRS IEKARSAAAK 201 EVQKMKNFGQ GGSQRIICKW ARMPNPGARK GSVPNWQSWAYLPKWSAIRR 251 DIKRFTACKA AICPPMR*

After further analysis, the complete gonococcal DNA sequence <SEQ ID389> was found to be:

1 ATGTTTATGA ACAAATTTTC CCAATCCGGA AAAGGTCTGT CCGGTTTCTT 51 CTTCGGTTTGATACTGGCAA CGGTCATTAT TGCCGGTATT TTGCTTTATC 101 TGAACCAGGG CGGTCAAAATGCGTTCAAAA TCCCGGCTCC GTCGAAGCAG 151 CCTGCAGAAA CGGAAATCCT GAAACTGAAAAACCAGCCTA AGGAAGACAT 201 CCAACCTGAA CCGGCCGATC AAAACGCCTT GTCCGAACCGGATGTTGCGA 251 AAGAGGCAGA GCAGTCGGAT GCGGAAAAAG CTGCCGACAA GCAGCCCGTT301 GCCGACAAag ccgacgAGGT TGAAGAAAag GcGGgcgAgc cggaACGGga 351aGAGCCGGAC ggACAGGCAG TGCGCAAGAA AGCACTGAcg gAAGAgcGTG 401 AACAAACcgtcagggAAAAA GCGCagaaga AAGATGCCGA AACGgTTAAA 451 AAacaaGCgg tAaaaccgtctAAAGAAACa gagaaaaaag cTtcaaaaga 501 agagaaaaag gcggcgaaag aaaAAGttgcacccaaaccg accccggaaC 551 aaatcctcaa cagccgCagc atcgaaaaag cgcgtagtgccgctgccaaa 601 gaAgtgcaGA AAatgaaaaa ctTtgggcaa ggcgGaagcc aacgcattaT651 CTGcaaatgg gcgcgtatgc cgaccgtccg gagcgcggaA gggcagcgtg 701ccaaACtggc aAtcttgGgc atatctTccg aagtggtcgG CTATCAGGCG 751 GGACATAAAACGCTTTACCG CGTGCAAagc GGCAatatgt ccgccgatgc 801 gGTGAAAAAA ATGCAGGACGAGTTGAAAAA GCATGGGGtt gcCAGCCTGA 851 TCCGTGcgAT TGAAGGCAAA TAA

This encodes the following amino acid sequence <SEQ ID 390>:

1 MFMNKFSQSG KGLSGFFFGL ILATVIIAGI LLYLNQGGQN AFKIPAPSKQ 51 PAETEILKLKNQPKEDIQPE PADQNALSEP DVAKEAEQSD AEKAADKQPV 101 ADKADEVEEK AGEPEREEPDGQAVRKKALT EEREQTVREK AQKKDAETVK 151 KQAVKPSKET EKKASKEEKK AAKEKVAPKPTPEQILNSRS IEKARSAAAK 201 EVQKMKNFGQ GGSQRIICKW ARMPTVRSAE GQRAKLAILGISSEVVGYQA 251 GHKTLYRVQS GNMSADAVKK MQDELKKHGV ASLIRAIEGK *

ORF65ng-1 and ORF65-1 show 89.0% identity in 290 aa overlap:

On this basis, including the presence of a putative transmembrane domainin the gonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 46

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 391>:

1 ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTACTCG GTkTCTTCGG 51 CGGAAcGCACTGCATCGGTA TGTGCGGCGG ATTAAGCAGC GcGTTTGs.s 101 TCCAACTCCC CCCGCATATCAACCGCTTTT GGCTGATCCT GCTGCTTAAC 151 ACAGGACGGG TAAGCAGCTA TACGGCAAtCGGCCTGATAC TCGGATTAAT 201 CGGACAGGTC GGCGTTTCAC TCGAcCAaAC CCGCGTCCTGCAGAATATTT 251 TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT ATACTTGAGC301 GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAaATCGGCA AACCGATATG 351GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA AAATCCATAC 401 CCGCCTGCCTtGCGgTCGGA ATATTATGGG GCTGGCTGCC GTGCGGACTG 451 GTTTACAGCG CGTCGCTTTACGCGCTGGGA AgCGGTAGTG CGGCAACGGG 501 CGGGTTATAT ATGCTTGCCT TTGCACTGGGTACGCTGCCC AATCTTtTAG 551 CAATCGGCAT TTTtTCCCTG CAACTGAAwA AAATCATGCAAAACCGATAT 601 ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT TATGGAAACT651 TGCCGTCCTG TGGCTGTAA

This corresponds to the amino acid sequence <SEQ ID 392; ORF103>:

1 MNHDITFLTL FLLGXFGGTH CIGMCGGLSS AFXXQLPPHI NRFWLILLLN 51 TGRVSSYTAIGLILGLIGQV GVSLDQTRVL QNILYTAANL LLLFLGLYLS 101 GISSLAAKIE KIGKPIWRNLNPILNRLLPI KSIPACLAVG ILWGWLPCGL 151 VYSASLYALG SGSAATGGLY MLAFALGTLPNLLAIGIFSL QLXKIMQNRY 201 IRLCTGLSVS LWALWKLAVL WL*

Further work elaborated the DNA sequence <SEQ ID 393> as:

1 ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTACTCG GTTTCTTCGG 51 CGGAACGCACTGCATCGGTA TGTGCGGCGG ATTAAGCAGC GCGTTTGCGC 101 TCCAACTCCC CCCGCATATCAACCGCTTTT GGCTGATCCT GCTGCTTAAC 151 ACAGGACGGG TAAGCAGCTA TACGGCAATCGGCCTGATAC TCGGATTAAT 201 CGGACAGGTC GGCGTTTCAC TCGACCAAAC CCGCGTCCTGCAGAATATTT 251 TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT ATACTTGAGC301 GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA AACCGATATG 351GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA AAATCCATAC 401 CCGCCTGCCTTGCGGTCGGA ATATTATGGG GCTGGCTGCC GTGCGGACTG 451 GTTTACAGCG CGTCGCTTTACGCGCTGGGA AGCGGTAGTG CGGCAACGGG 501 CGGGTTATAT ATGCTTGCCT TTGCACTGGGTACGCTGCCC AATCTTTTAG 551 CAATCGGCAT TTTTTCCCTG CAACTGAAAA AAATCATGCAAAACCGATAT 601 ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT TATGGAAACT651 TGCCGTCCTG TGGCTGTAA

This corresponds to the amino acid sequence <SEQ ID 394; ORF103-1>:

1 MNHDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI NRFWLILLLN 51TGRVSSYTAI GLILGLIGQV GVSLDQTRVL QNILYTAANL LLLFLGLYLS 101 GISSLAAKIEKIGKPIWRNL NPILNRLLPI KSIPACLAVG ILWGWLPCGL 151 VYSASLYALG SGSAATGGLYMLAFALGTLP NLLAIGIFSL QLKKIMQNRY 201 IRLCTGLSVS LWALWKLAVL WL*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF103 shows 93.8% identity over. a 222aa overlap with an ORF (ORF103a)from strain A of N. meningitidis:

The complete length ORF103a nucleotide sequence <SEQ ID 395> is:

1 ATGAACCANG ACATCACTTT CCTCACCCTG TTCCTACTCG GTTTCTTCGG 51 CGGAACGCACTGCATCGGTA TGTGCGGCGG ATTAAGCAGC GCGTTTGCGC 101 TCCAACTCCC CCCGCATATCAACCGCTTNT GGCTGATCCT GCTGCTTAAC 151 ACAGGACGGG TAAGCAGCTA TACGGCAATCGGCCTGATAC TCGGATTAAT 201 CGGACAGGTC GGCGTTTCAC TCGACCAAAC CCGCGTCNTGCAGAATATTT 251 TATACACGGC CGCCAACCTC CTGCTGCTCT TTTTAGGCTT ATACTTGAGC301 GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA AACCGATATG 351GCGGAACCTG AACCCGATAC TCAACCGGCT GTTACCCATA AAATCCATAC 401 CCGCCTGCCTTGCGGTCGGA ATATTATGGG GCTGGCTGCC GTGCGGACTA 451 GTTTACAGCG CGTCGCTTTACGCGCTGGGA AGCGGTAGTG CGGCAACGGG 501 CGGGTTATAT ATGCTTGCCT TTGCACTGGGTACGCTGCCC AATCTTTNGG 551 CAATCGGCAT TTTTTCCCTG CAACTGNAAA AAATCATGCAAAACCGATAT 601 ATCCGCCTGT GTACGGGATT ATCCGTATCA TTATGGGCAT TATGGAAACT651 TGCCGTCCTG TGGCTGTAA

This encodes a protein having amino acid sequence <SEQ ID 396>:

1 MNXDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI NRXWLILLLN 51TGRVSSYTAI GLILGLIGQV GVSLDQTRVX QNILYTAANL LLLFLGLYLS 101 GISSLAAKIEKIGKPIWRNL NPILNRLLPI KSIPACLAVG ILWGWLPCGL 151 VYSASLYALG SGSAATGGLYMLAFALGTLP NLXAIGIFSL QLXKIMQNRY 201 IRLCTGLSVS LWALWKLAVL WL*

ORF103a and ORF103-1 show 97.7% identity in 222 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF103 shows 95.5% identity over a 222aa overlap with a predicted ORF(ORF103.ng) from N. gonorrhoeae:

The complete length ORF103ng nucleotide sequence <SEQ ID 397> is:

1 ATGAACCACG ACATCACTTT CCTCACCCTG TTCCTGCTCG GTTTCTTCGG 51 CGGAACTCACTGCATCGGTA TGTGCGGCGG ATTAAGCAGC GCGTTTGCGC 101 TCCAACTCCC CCCGCATATCAACCGCTTTT GGCTGATTCT GCTGCTTAAC 151 ACAGGACGGA TAAGCAGCTA TACGGCAATCGGCCTGATGC TCGGATTAAT 201 CGGACAACTC GGCATTTCAC TCGACCAAAc ccgcgTCCTGCAAAATATTT 251 tatacacagc ctccaaCCTC CTGCTGCTCT TTTTAGGCTT ATACTTGAGC301 GGTATTTCTT CCTTGGCGGC AAAAATCGAG AAAATCGGCA AACCGATATG 351GCGCAACCTG AACCCGATAC TCAACCGGCT GCTGCCCATA AAATCCATAC 401 CCGCCTGCCTTGCTGTCGGA ATATTATGGG GCTGGCTGCC GTGCGGACTG 451 GTTTACAGCG CATCACTTTACGCGCTGGGA AGCGGTAGTG CGACAACCGG 501 CGGACTGTAT ATGCTTGCCT TTGCACTGGGTACGCTGCCC AATCTTTTGG 551 CAATCGGCAT TTTTTCCCTG CAACTGAAAA AAATCATGCAAAACCGATAT 601 ATCCGCCTGT GTACAGGATT ATCCGTATCA TTATGGGCAT TATGGAAGCT651 TGCCGTCCTG TGGCTGTAA

This encodes a protein having amino acid sequence <SEQ ID 398>:

1 MNHDITFLTL FLLGFFGGTH CIGMCGGLSS AFALQLPPHI NRFWLILLLN 51TGRISSYTAI GLMLGLIGQL GISLDQTRVL QNILYTASNL LLLFLGLYLS 101 GISSLAAKIEKIGKPIWRNL NPILNRLLPI KSIPACLAVG ILWGWLPCGL 151 VYSASLYALG SGSATTGGLYMLAFALGTLP NLLAIGIFSL QLKKIMQNRY 201 IRLCTGLSVS LWALWKLAVL WL*

In addition, ORF103ng and ORF103-1 show 97.3% identity in 222 aaoverlap:

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 47

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 399>:

1 ATGGAAAACC AAAGGCCGCT CCTAGGCTTT CGCTTGGCAC TTTTGGCGGC 51 GATGACGTGGGGAACGCTGC CGAT.TCCGT GCGGCAGGTA TTGAAGTTTG 101 TCGATGCGCC GACGCTGGTGTGGGTGCGTT TTACCGTGGC GGCGGCGGTA 151 TTGTTTGTTT TGCTGGCACT GGGCGGGCGGCTGCcGAAGC GGCGaGGATT 201 TTTCTTGGTG CTCATTCAGG CTGCTGCTGC TCGGCGTGGCGGGCATTTCG 251 GCAAACTTTG TGCTGATTGC CCAAGGGCTG CATTATATTT CGCCGACCAC301 GACGCAGGTT TTGTGGCAGA TTTCGCCGTT TACGATGATT GTwGTCGGTG 351TGTTGGTGTT TAAAGACCGG ATGACTGCCG CTCAGAAAAT CGGCTTGGTT 401 TTGCTGCTTGCCGGTTTGCT TATGTATTTT AACGATAAAT TCGGCGAGTT 451 GTCGGGTTTG GGCGCGTATGC.AAGGGCGT GTTGCTGTGT GCGGCAGGCA 501 GTATGGCATG GGTGTGTAAT GCCGTGGCGCAAAAGCTGCT GTCGGCGCAA 551 TTCGGGCCGC AACAGATTCT GCTGTTGATT TATGCGGCAAGTGCCGCCGT 601 GTTCCTGCCG TTTGCCGAAC CGGCACACAT CGGAAGTATG GACGGTACGT651 TGGCGTGGGT ATGTATTGCG TATTGCTGCT TGAATACGTT AATCGGTTAC 701GGCTCGTTCG GCGAGGCGTT GAAACATTGG GAGGCTTCCA AAGTCAGCGC 751 GGTAACAACCTTGCTCCCCG TGTTTACCGT AATAAATACT TTGCTCGGGC 801 ATTATGTGAT GCCTGAAACTTTTGCCGCGC CGGA..

This corresponds to the amino acid sequence <SEQ ID 400; ORF104>:

1 MENQRPLLGF RLALLAAMTW GTLPXSVRQV LKFVDAPTLV WVRFTVAAAV 51 LFVLLALGGRLPKRRDFSWC SFRLLLLGVA GISANFVLIA QGLHYISPTT 101 TQVLWQISPF TMIVVGVLVFKDRMTAAQKI GLVLLLAGLL MYFNDKFGEL 151 SGLGAYXKGV LLCAAGSMAW VCNAVAQKLLSAQFGPQQIL LLIYAASAAV 201 FLPFAEPAHI GSMDGTLAWV CIAYCCLNTL IGYGSFGEALKHWEASKVSA 251 VTTLLPVFTV INTLLGHYVM PETFAAP...

Further work revealed further, partial DNA sequence <SEQ ID 401>:

1 ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC TTTTGGCGGC 51 GATGACGTGGGGAACGCTGC CGATTGCCGT GCGGCAGGTA TTGAAGTTTG 101 TCGATGCGCC GACGCTGGTGTGGGTGCGTT TTACCGTGGC GGCGGCGGTA 151 TTGTTTGTTT TGCTGGCACT GGGCGGGCGGCTGCCGAAGC GGCGGGATTT 201 TTCTTGGTGC TCATTCAGGC TGCTGCTGCT CGGCGTGGCGGGCATTTCGG 251 CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC GCCGACCACG301 ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG TTGTCGGTGT 351GTTGGTGTTT AAAGACCGGA TGACTGCCGC TCAGAAAATC GGCTTGGTTT 401 TGCTGCTTGCCGGTTTGCTT ATGTTTTTTA ACGATAAATT CGGCGAGTTG 451 TCGGGTTTGG GCGCGTATGCGAAGGGCGTG TTGCTGTGTG CGGCAGGCAG 501 TATGGCATGG GTGTGTTATG CCGTGGCGCAAAAGCTGCTG TCGGCGCAAT 551 TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGCAAGTGCCGCCGTG 601 TTCCTGCCGT TTGCCGAACC GGCACACATC GGAAGTTTGG ACGGTACGTT651 GGCGTGGGTT TGTTTTGCGT ATTGCTGCTT GAATACGTTA ATCGGTTACG 701GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA AGTCAGCGCG 751 GTAACAACCTTGCTCCCCGT GTTTACCGTA ATAwTwwCTT TGCTCGGGCA 801 TTATGTGATG CCTGAAACTTTTGCCGCGCC GGA...

This corresponds to the amino acid sequence <SEQ ID 402; ORF104-1>:

1 MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV WVRFTVAAAV 51 LFVLLALGGRLPKRRDFSWC SFRLLLLGVA GISANFVLIA QGLHYISPTT 101 TQVLWQISPF TMIVVGVLVFKDRMTAAQKI GLVLLLAGLL MFFNDKFGEL 151 SGLGAYAKGV LLCAAGSMAW VCYAVAQKLLSAQFGPQQIL LLIYAASAAV 201 FLPFAEPAHI GSLDGTLAWV CFAYCCLNTL IGYGSFGEALKHWEASKVSA 251 VTTLLPVFTV IXXLLGHYVM PETFAAP...

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical HI0878 Protein of H. influenzae (AccessionNumber U32769)

ORF104 and HI0878 show 40% aa identity in 277aa overlap:

orf104 4 QRPLLGFRLALLAAMTWGTLPXSVRQVLKFVDAPTLVWXXXXXXXXXXXXXXXXXXXXP- 62Q+PLLGF  AL+ AM WG+LP +++QVL  ++A T+VW                    P HI0878 3QQPLLGFTFALITAMAWGSLPIALKQVLSVMNAQTIVWYRFIIAAVSLLALLAYKKQLPE 62 orf10463 --KRRDFSWCSFRLLLLGVAGISANFVLIAQGLHYISPTTTQVLWQISPFTMIVVGVLVF 120   KR ++W    ++L+GV G+++NF+L +  L+YI P+  Q+   +S F M++ GVL+F HI0878 63LMKVRQYAW----IMLIGVIGLTSNFLLFSSSLNYIEPSVAQIFIHLSSFGMLICGVLIF 118 orf104121 KDRMTAAQKIXXXXXXXXXXMYFNDKFGELSGLGAYXKGVLLCAAGSMAWVCNAVAQKLL 180K+++   QKI          ++FND+F   +GL  Y  GV+L   G++ WV   +AQKL+ HI0878 119KEKLGLHQKIGLFLLLIGLGLFFNDRFDAFAGLNQYSTGVILGVGGALIWVAYGMAQKLM 178 orf104181 SAQFGPQQILLLIYAASAAVFLPFAEPAHIGSMDGTLAWVCIAYCCLNTLIGYGSFGEAL 240  +F  QQILL++Y   A  F+P A+ + +  +   LA +C  YCCLNTLIGYGS+ EAL HI0878 179LRKFNSQQILLMMYLGCAIAFMPMADFSQVQELT-PLALICFIYCCLNTLIGYGSYAEAL 237 orf104241 KHWEASKVSAVTTLLPVFTVINTLLGHYVMPETFAAP 277   W+ SKVS VTL+P+FT++ + + HY  P  FAAP HI0878 238NRWDVSKVSVVITLVPLFTILFSHIAHYFSPADFAAP 274Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF104 shows 95.3% identity over a 277aa overlap with an ORF (ORF104a)from strain A of N. meningitidis:

The complete length ORF104a nucleotide sequence <SEQ ID 403> is:

1 ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC TTTTGGCGGC 51 GATGACGTGGGGAACGCTGC CGATTGCCGT GCGGCAGGTA TTGAAGTTTG 101 TCGATGCGCC GACGCTGGTGTGGGTGCGTT TTACCGTGGC GGCGGCGGTA 151 TTGTTTGTTT TGCTGGCATT GGGCGGGCGGCTGCCGAAGT GGCGGGATTT 201 TTCTTGGTGC TCATTCAGGC TGCTGCTGCT CGGCGTGGCGGGCATTTCGG 251 CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC GCCGACCACG301 ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG TTGTCGGTGT 351GTTGGTGTTT AAAGACCGGA TGACTGCCGC TCAGAAAATC GGCTTGGTTT 401 TGCTGCTTGCCGGTTTGCTT ATGTTTTTTA ACGATAAATT CGGCGAGTTG 451 TCGGGTTTGG GCGCGTATGCGAAGGGCGTG TTGCTGTGTG CGGCAGGCAG 501 TATGGCATGG GTGTGTTATG CCGTGGCGCAAAAGCTGCTG TCGGCGCAAT 551 TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGCAAGTGCCGCCGTG 601 TTCCTGCCGT TTGCCGAACT GGCACACATC GGAAGTTTGG ACGGTACGTT651 GGCGTGGGTT TGTTTTGCGT ATTGCTGCTT GAATACGTTA ATCGGTTACG 701GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA AGTCAGCGCG 751 GTAACAACCTTGCTCCCCGT GTTTACCGTA ATATTTTCTT TGCTCGGGCA 801 TTATGTGATG CCTGATACTTTTGCCGCGCC GGATATGAAC GGTTTGGGTT 851 ATGCCGGCGC ACTGGTCGTG GTCGGGGGTGCGGTTACGGC GGCGGTGGGG 901 GACAGGCTGT TCAAACGCCG CTAG

This encodes a protein having amino acid sequence <SEQ ID 404>:

1 MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV WVRFTVAAAV 51 LFVLLALGGRLPKWRDFSWC SFRLLLLGVA GISANFVLIA QGLHYISPTT 101 TQVLWQISPF TMIVVGVLVFKDRMTAAQKI GLVLLLAGLL MFFNDKFGEL 151 SGLGAYAKGV LLCAAGSMAW VCYAVAQKLLSAQFGPQQIL LLIYAASAAV 201 FLPFAELAHI GSLDGTLAWV CFAYCCLNTL IGYGSFGEALKHWEASKVSA 251 VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYAGALVV VGGAVTAAVG301 DRLFKRR*

ORF104a and ORF104-1 show 98.2% identity in 277 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF104 shows 93.9% identity over a 277aa overlap with a predicted ORF(ORF104.ng) from N. gonorrhoeae:

The complete length ORF104ng nucleotide sequence <SEQ ID 405> ispredicted to encode a protein having amino acid sequence <SEQ ID 406>:

1 MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV WVRFTVAAAV 51 LFVLLALGGRLPKRRDFSWH SFRLLLLGVT GISANFVLIA QGLHYISPTT 101 TQVLWQISPF TMIVVGVLVFKDRMTAAQKI GLVLLLVGLL MFFNDKFGEL 151 SGLGAYAKGV LLCAAGSMAW VCYAVAQKLLSAQFGPQQIL LLIYAASAAV 201 FLLXAEPAHI GSLDGTLAWV CFVYCCLNTL IGYGSFGEALKHWEASKVSA 251 VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYVGALVV VGGAVTAAVG301 DRPFKRR*

Further work revealed the complete gonococcal nucleotide sequence <SEQID 407>:

1 ATGGAAAACC AAAGGCCGCT CCTAGGCTTC GCGTTGGCAC TTTTGGCGGC 51 GATGACGTGGGGGACGCTGC CGATTGCCGT GCGGCAGGTA TTGAAGTTTG 101 TCGATGCGCC GACGCTGGTGTGGGTGCGTT TTACCGTGGC GGCGGCGGTA 151 TTGTTTGTTT TGCTGGCATT GGGCGGGCGGCTGCCGAAGC GGCGGGATTT 201 TTCTTGGCAT TCATTCAGGC TGCTGCTGCT CGGCGTGACGGGCATTTCGG 251 CAAACTTTGT GCTGATTGCC CAAGGGCTGC ATTATATTTC GCCGACCACG301 ACGCAGGTTT TGTGGCAGAT TTCGCCGTTT ACGATGATTG TTGTCGGCGT 351GTTGGTGTTT AAAGACCGGA tgaCTGCCGC GCAGAAAATC GGTTTGGTTT 401 TGCTGCttgTCGGTttgCTT ATGTTTTtta ACGACAAATT CGGCGAGTTG 451 TCGGGTTTGG GCGCGTATGCGAAGGGCGTG TTGCTGTGTG CGGCAGGCAG 501 TATGGCCTGG GTGTGTTATG CCGTGGCGCAAAAGCTGCTG TCGGCGCAAT 551 TCGGGCCGCA ACAGATTCTG CTGTTGATTT ATGCGGcaagtgccgccGTG 601 TTCCtgccgT TTGccgaaCC GGCACACATC GGAAGTTTgg aCGGTACGtt651 GGCGTGGGTT TGTTTTGTGT ATTGCTGCTT GAATACGTTA ATCGGTTACG 701GCTCGTTCGG CGAGGCGTTG AAACATTGGG AGGCTTCCAA AGTCAGCGCG 751 GTAACAACCTTGCTCCCCGT GTTTACCGTA ATATTTTCTT TGCTCGGGCA 801 TTATGTGATG CCTGATACTTTTGCCGCGCC GGATATGAAC GGTTTGGGTT 851 ATGTCGGCGC ACTGGTCGTG GTCGGGGGTGCGGTTACGGC GGCGGTGGGG 901 GACAGGCCGT TCAAACGCCG CTAG

This corresponds to the amino acid sequence <SEQ ID 408; ORF104ng-1>:

1 MENQRPLLGF ALALLAAMTW GTLPIAVRQV LKFVDAPTLV WVRFTVAAAV 51 LFVLLALGGRLPKRRDFSWH SFRLLLLGVT GISANFVLIA QGLHYISPTT 101 TQVLWQISPF TMIVVGVLVFKDRMTAAQKI GLVLLLVGLL MFFNDKFGEL 151 SGLGAYAKGV LLCAAGSMAW VCYAVAQKLLSAQFGPQQIL LLIYAASAAV 201 FLPFAEPAHI GSLDGTLAWV CFVYCCLNTL IGYGSFGEALKHWEASKVSA 251 VTTLLPVFTV IFSLLGHYVM PDTFAAPDMN GLGYVGALVV VGGAVTAAVG301 DRPFKRR*

ORF104ng-1 and ORF104-1 show 97.5% identity in 277 aa overlap:

In addition, ORF104ng-1 shows significant homology with a hypotheticalH. influenzae protein:

gi|1573895 (U32769) hypothetical [Haemophilus influenzae] Length = 306Score = 237 bits (598), Expect = 8e−62 Identities = 114/280 (40%),Positives = 168/280 (59%), Gaps = 8/280 (2%) Query: 30QRPXXXXXXXXXXXMTWGTLPIAVRQVLKFVDAPTLVWXXXXXXXXXXXXXXXXXXXXP- 88Q+P           M WG+LPIA++QVL  ++A T+VW                    P Sbjct: 3QQPLLGFTFALITAMAWGSLPIALKQVLSVMNAQTIVWYRFIIAAVSLLALLAYKKQLPE 62 Query:89 --KRRDFSWHSFRLLLLGVTGISANFVLIAQGLHYISPTTTQVLWQISPFTMIVVGVLVF 146   KR ++W    ++L+GV G+++NF+L +  L+YI P+  Q+   +S F M++ GVL+F Sbjct: 63LMKVRQYAW----IMLIGVIGLTSNFLLFSSSLNYIEPSVAQIFIHLSSFGMLICGVLIF 118 Query:147 KDRMTAAQKIXXXXXXXXXXMFFNDKFGELSGLGAYAKGVLLCAAGSMAWVCYAVAQKLL 206K+++   QKI          +FFND+F   +GL  Y+ GV+L   G++ WV Y +AQKL+ Sbjct: 119KEKLGLHQKIGLFLLLIGLGLFFNDRFDAFAGLNQYSTGVILGVGGALIWVAYGMAQKLM 178 Query:207 SAQFGPQQILLLIYAASAAVFLPFAEPAHIGSLDGTLAWVCFVYCCLNTLIGYGSFGEAL 266  +F  QQILL++Y   A  F+P A+ + +  L   LA +CF+YCCLNTLIGYGS+ EAL Sbjct: 179LRKFNSQQILLMMYLGCAIAFMPMADFSQVQELT-PLALICFIYCCLNTLIGYGSYAEAL 237 Query:267 KHWEASKVSAVTTLLPVFTVIFSLLGHYVMPDTFAAPDMN 306   W+ SKVS V TL+P+FT++FS+ HY  P  FAAP++N Sbjct: 238 NRWDVSKVSVVITLVPLFTILFSHIAHYFSPADFAAPELN 277

Based on this analysis, including the presence of a putative leadersequence and several putative transmembrane domains in the gonococcalprotein, it is predicted that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 48

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 409>:

1 ATGGTAGCTC GTCGGGCTCA TAACCCGAAG GTCGTAGGTT CGAATCCTGT 51 .CCCGCAACCTAATTTCAAA CCCCTCGGTT CAATGCCGAG GG.GTTTTGT 101 T.TTGCCTGT TTCCTGTTTCCTGTTTCCTG CCGCCTCCGT TTTTTGCCGG 151 ATTTTCCTTC CGGCCGCAAT ATCGGAACGGCAGACCGCCG TCTGTTTGCG 201 GTTGCAAATT CAGGCAGTTT GGCTACAATC TTCCGCATTGTCTTCAAGAA 251 AGCCAACCAT GCCGACCGTC CGTTTTACCG AATCCGTCAG CAAACAAGAC301 CTTGATGCTC TGTTCGAGTG GGCAAAAGCA AGTTACGGTG CAGAAAGTTG 351CTGGAAAACG CTGTATCTGA ACGGTCysCC TTTGGGCAAC CTGTCGCCGG 401 AATGGGTGGAACGCGTsmmA AAAGACTGGG AGGCAGGCTG CyCGGAGTCT 451 TCAGACGGCA TTTTTCTGAATgCGGACGGc TGgCctGATA TGGgCGGAcg 501 cTTACAGCAC CTCGCCCTCG GTTGGCACTGTGCGGGGCTG TTGGACGgsT 551 GGCGCAACGA GTGTTTCGAC CTGACCGACG GCGGCGGCAACCCCTTGTTC 601 ACGCTCGaAc GCGCCGyTTT mCGTCCTkTC GGACTGCTCA GCCGCGCCGT651 CCATCTCAAC GGTCTGACCG AATCGGACGG CCGATGGCAT TTCTGGATAG 701GCAGGCGCAG TCCGCACAAA GCAGTCGATC CCAACAAACT CGACAATACT 751 rCCGCCGGCGGTGTTTCCGG CGGCGAAATG CCGTCTGAAG CCGTGTGTCG 801 CGAAAGCAGC GAAGAAGCCGGTTTGGATAA AACGCTGcTT CCGCTCATCC 851 GCCCGGTATC GCAGCTGCAC AGCCTGCGCTCCGTCAGCCG GGGTGTACAC 901 AATGAAATCC TGTATGTATT CGATGCCGTC CTGCCG...

This corresponds to the amino acid sequence <SEQ ID 410; ORF105>:

1 MVARRAHNPK VVGSNPXPAT XFQTPRFNAE XVLXLPVSCF LFPAASVFCR 51 IFLPAAISERQTAVCLRLQI QAVWLQSSAL SSRKPTMPTV RFTESVSKQD 101 LDALFEWAKA SYGAESCWKTLYLNGXPLGN LSPEWVERVX KDWEAGCXES 151 SDGIFLNADG WPDMGGRLQH LALGWHCAGLLDGWRNECFD LTDGGGNPLF 201 TLERAXXRPX GLLSRAVHLN GLTESDGRWH FWIGRRSPHKAVDPNKLDNT 251 XAGGVSGGEM PSEAVCRESS EEAGLDKTLL PLIRPVSQLH SLRSVSRGVH301 NEILYVFDAV LP...

Further work revealed the complete nucleotide sequence <SEQ ID 411>:

1 ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACAAG ACCTTGATGC 51 TCTGTTCGAGTGGGCAAAAG CAAGTTACGG TGCAGAAAGT TGCTGGAAAA 101 CGCTGTATCT GAACGGTCTGCCTTTGGGCA ACCTGTCGCC GGAATGGGTG 151 GAACGCGTCA AAAAAGACTG GGAGGCAGGCTGCTCGGAGT CTTCAGACGG 201 CATTTTTCTG AATGCGGACG GCTGGCCTGA TATGGGCGGACGCTTACAGC 251 ACCTCGCCCT CGGTTGGCAC TGTGCGGGGC TGTTGGACGG CTGGCGCAAC301 GAGTGTTTCG ACCTGACCGA CGGCGGCGGC AACCCCTTGT TCACGCTCGA 351ACGCGCCGCT TTCCGTCCTT TCGGACTGCT CAGCCGCGCC GTCCATCTCA 401 ACGGTCTGACCGAATCGGAC GGCCGATGGC ATTTCTGGAT AGGCAGGCGC 451 AGTCCGCACA AAGCAGTCGATCCCAACAAA CTCGACAATA CTGCCGCCGG 501 CGGTGTTTCC GGCGGCGAAA TGCCGTCTGAAGCCGTGTGT CGCGAAAGCA 551 GCGAAGAAGC CGGTTTGGAT AAAACGCTGC TTCCGCTCATCCGCCCGGTA 601 TCGCAGCTGC ACAGCCTGCG CTCCGTCAGC CGGGGTGTAC ACAATGAAAT651 CCTGTATGTA TTCGATGCCG TCCTGCCCGA AACCTTCCTG CCTGAAAATC 701AGGATGGCGA AGTGGCGGGT TTTGAGAAAA TGGACATCGG CGGTCTGTTG 751 GATGCCATGTTGTCGGGAAA CATGATGCAC GACGCGCAAC TGGTTACGCT 801 GGACGCGTTT TGCCGTTACGGTCTGATTGA TGCCGCCCAT CCGCTGTCCG 851 AGTGGCTGGA CGGCATACGT TTATAG

This corresponds to the amino acid sequence <SEQ ID 412; ORF105-1>:

1 MPTVRFTESV SKQDLDALFE WAKASYGAES CWKTLYLNGL PLGNLSPEWV 51 ERVKKDWEAGCSESSDGIFL NADGWPDMGG RLQHLALGWH CAGLLDGWRN 101 ECFDLTDGGG NPLFTLERAAFRPFGLLSRA VHLNGLTESD GRWHFWIGRR 151 SPHKAVDPNK LDNTAAGGVS GGEMPSEAVCRESSEEAGLD KTLLPLIRPV 201 SQLHSLRSVS RGVHNEILYV FDAVLPETFL PENQDGEVAGFEKMDIGGLL 251 DAMLSGNMMH DAQLVTLDAF CRYGLIDAAH PLSEWLDGIR L*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF105 shows 89.4% identity over a 226aa overlap with an ORF (ORF105a)from strain A of N. meningitidis:

The complete length ORF105a nucleotide sequence <SEQ ID 413> is:

1 ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACACG ACCTTGATGC 51 CCTATTCGAGTGGGCAAAGG CAAGTTACGG TGCGGAAAGT TGCTGGAAAA 101 CGCTGTATCT GAACGGTCTGCCTTTGGGCA ATCTGTCGCC GGAATGGGCG 151 GAGCGCGTCA AAAAAGACTG GGAGGCAGGCTGCTCGGAGT CTTCAGACGG 201 CATTTTCCTG AATGCGGACG GCTGGCCAGA TATGGGCAGACGCTTGCAGC 251 ACCTCGCCCG AATATGGAAA GAAGCGGGAC TGCTTCACGG CTGGCGCGAC301 GAGTGTTTCG ACCTGACCGA CGGCGGCAGC AATCCCTTGT TCGCGCTCGA 351ACGCGCCGCT TTCCGTCCGT TCGGACTGCT CAGCCGCGCC GTCCATCTCA 401 ACGGTTTGGTCGAATCGGAC GGCCGATGGC ATTTCTGGAT AGGCAGGCGC 451 AGTCCGCACA AAGCAGTCGATCCCGACAAA CTCGACAATA CTGCCGCCGG 501 CGGTGTTTCC AGCGGTGAAT TGCCGTCTGAAACCGTGTGT CGCGAAAGCA 551 GCGAAGAAGC CGGTTTGGAT AAAACGCTGC TTCCGCTCATCCGCCCGGTA 601 TCGCAGCTGC ACAGCCTGCG CCCCGTCAGC CGGGGTGTGC ACAATGAAAT651 CCTGTATGTA TTCGATGCCG TCCTGCCCGA AACCTTCCTG CCTGAAAATC 701AGGATGGCGA AGTGGCGGGT TTTGAGAAAA TGGACATCGG CGGTCTGTTG 751 GCTGCCATGTTGTCGGGAAA CATGATGCAC GACGCGCAAC TGGTTACGCT 801 GGACGCGTTT TGCCGTTACGGTCTGATTGA TGCCGCCCAT CCGCTGTCCG 851 AGTGGCTGGA CGGCATACGT TTATAG

This encodes a protein having amino acid sequence <SEQ ID 414>:

1 MPTVRFTESV SKHDLDALFE WAKASYGAES CWKTLYLNGL PLGNLSPEWA 51 ERVKKDWEAGCSESSDGIFL NADGWPDMGR RLQHLARIWK EAGLLHGWRD 101 ECFDLTDGGS NPLFALERAAFRPFGLLSRA VHLNGLVESD GRWHFWIGRR 151 SPHKAVDPDK LDNTAAGGVS SGELPSETVCRESSEEAGLD KTLLPLIRPV 201 SQLHSLRPVS RGVHNEILYV FDAVLPETFL PENQDGEVAGFEKMDIGGLL 251 AAMLSGNMMH DAQLVTLDAF CRYGLIDAAH PLSEWLDGIR L*

ORF105a and ORF105-1 show 93.8% identity in 291 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF105 shows 87.5% identity over a 312aa overlap with a predicted ORF(ORF105.ng) from N. gonorrhoeae:

A complete length ORF105ng nucleotide sequence <SEQ ID 415> waspredicted to encode a protein having amino acid sequence <SEQ ID 416>:

1 MVARRAHNPK VVGSNPAPAT KYQTPRFNAE GVLFFLFPAA SVFCRIFLPA 51 AISERQAAVCLRLQIQAVWL QSSALCSRKP AMPTVRFTES VSKQDLDALF 101 ERAKASYGAE SCWKTLYLNRLPLGNLSPEW AERIKKDWEA GCSESSNGIF 151 LNADGWPDMG GRLQHLARTW NKAGLLHGWRNECFDLTDGG GNPLFTLERA 201 AFRPFGLLIR AVHLNGLVES NGRWHFWIGR RSPHKAVDPGKLDNIAGGGV 251 SGGEMPSEAV CRESSEEAGL DKTLFPLIRP VSRLHSLRPV SRGVHNEILY301 VFDAVLPETF LPENQDGEVA GFEKMDIGGL LDAMLSKNMM HDAQLVTLDA 351FYRYGLIDAA HPLSEWLDGI RL*

Further work revealed the complete nucleotide sequence <SEQ ID 417>:

1 ATGCCGACCG TCCGTTTTAC CGAATCCGTC AGCAAACAAG ACCTTGATGC 51 CCTGTTCGAGCGGGCAAAAG CAAGTTACGG TGCCGAAAGT TGCTGGAAAA 101 CGCTGTATCT GAACCGTCTTCCTTTGGGCA ATCTGTCGCC GGAATGGGCT 151 GAGCGCATCA AAAAAGACTG GGAGGCAGGCTGCTCCGAGT CTTCAGACGG 201 CATTTTTCTG AATGCGGACG GCTGGCCGGA TATGGGCGGACGCTTGCAGC 251 ACCTCGCCCG CACATGGAAC AAGGCGGGGC TGCTTCACGG ATGGCGCAAC301 GAGTGTTTCG ACCTGACCGA CGGCGGCGGC AACCCCTTGT TCACGCTCGA 351ACGCGCCGCT TTCCGTCCGT TCGGACTACT CAGCCGCGCC GTCCATCTCA 401 ACGGTTTGGTCGAATCGAAC GGCAGATGGC ATTTTTGGAT AGGCAGGCGC 451 AGTCCGCACA AAGCAGTCGatcCCGGCAAG CTCGACAATA TTGCCGGCGG 501 CGGTGTTTCC GGCGGCGAAA TGCCGTCTGAAGCCGTGTGC CGCGAAAGCA 551 GCGAAGAAGC CGGTTTGGAT AAAACGCTGT TTCCGCTCATCCGCCCAGTA 601 TCGCGGCTGC ACAGCCTTCG CCCCGTCAGC CGAGGTGTGC ACAATGAAAT651 CCTGTATGTG TTCGATGCCG TCCTGCCCGA AACCTTCCTG CCTGAAAATC 701AGGATGGCGA GGTAGCGGGT TTTGAAAAGA TGGACATTGG CGGCCTATTG 751 GATGCCATGTTGTCGAAAAA CATGATGCAC GACGCGCAAC TGGTTACGCT 801 GGACGCGTTT TACCGTTACGGTCTGATTGA TGCCGCCCAT CCGCTGTCCG 851 AGTGGCTGGA CGGCATACGT TTATAG

This corresponds to the amino acid sequence <SEQ ID 418; ORF105ng-1>:

1 MPTVRFTESV SKQDLDALFE RAKASYGAES CWKTLYLNRL PLGNLSPEWA 51 ERIKKDWEAGCSESSDGIFL NADGWPDMGG RLQHLARTWN KAGLLHGWRN 101 ECFDLTDGGG NPLFTLERAAFRPFGLLSRA VHLNGLVESN GRWHFWIGRR 151 SPHKAVDPGK LDNIAGGGVS GGEMPSEAVCRESSEEAGLD KTLFPLIRPV 201 SRLHSLRPVS RGVHNEILYV FDAVLPETFL PENQDGEVAGFEKMDIGGLL 251 DAMLSKNMMH DAQLVTLDAF YRYGLIDAAH PLSEWLDGIR L*

ORG105ng-1 and ORF105-1 show 93.5% identity in 291 aa overlap:

Furthermore, ORF105ng-1 shows homology with a yeast enzyme:

sp|P41888|TNR3_SCHPO THIAMIN PYROPHOSPHOKINASE (TPK) (THIAMINKINASE) >gi|1076928|pir||S52350 thiamin pyrophosphokinase (EC 2.7.6.2) -fission yeast (Schizosaccharomyces pombe) >gi|666111 (X84417) thiaminpyrophosphokinase [Schizosaccharomycespombe] >gi|2330852|gnl|PID|e334056 (Z98533) thiamin pyrophosphokinase[Schizosaccharomyces pombe] Length = 569 Score = 105 bits (259), Expect= 4e−22 Identities = 64/192 (33%), Positives = 94/192 (48%), Gaps= 3/192 (1%) Query: 268NKAGLLHGWRNECFDLTDGGGNPLFTLERAAFRPFGLLSRAVHLNGLVESNGRW--HFWI 441N  G+   WRNE + +      P+  +ER  F  FG LS  VH    + +        W+ Sbjct: 96NTFGIADQWRNELYTVYGKSKKPVLAVERGGFWLFGFLSTGVHCTMYIPATKEHPLRIWV 155 Query:442 GRRSPHKAVDPGKLDNIAGGGVSGGEMPSEAVCRESSEEAGLDKTLFPLIRPVSRLHSLR 621 RRSP K   P  LDN   GG++ G+     + +E SEEA LD +   LI P   +  ++ Sbjct: 156PRRSPTKQTWPNYLDNSVAGGIAHGDSVIGTMIKEFSEEANLDVSSMNLI-PCGTVSYIK 214 Query:622 PVSRG-VHNEILYVFDAVLPETFLPENQDGEVAGFEKMDIGGLLDAMLSKNMMHDAQLVT 798   R  +  E+ YVFD  + +  +P   DGEVAGF  + +  +L  +  K+   +  LV Sbjct: 215MEKRHWIQPELQYVFDLPVDDLVIPRINDGEVAGFSLLPLNQVLHELELKSFKPNCALVL 274 Query:799 LDAFYRYGLIDAAHP 843 LD   R+G+I   HP Sbjct: 275 LDFLIRHGIITPQHP 289

Based on this analysis, including the presence of a putativetransmembrane domain in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 49

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 419>:

1 ATGAATAGAC CCAAGCAACC CTTCTTCCGT CCCGAAGTCG CCGTTGCCCG 51 CCAAACCAGCCTGACGGGTA AAGTGATTCT GACACGACCG TTGTCATTTT 101 CCCTATGGAC GACATTTGCATCGATATCTG CGTTATTGAT TATCCTGTTT 151 TTGATATTTG GTAACTATAC GCGAAAGACAACAGTGGAGG GACAAATTTT 201 ACCTGCATCG GGCGTAATCA GGGTGTATGC ACCGgATACGrGkACAATTA 251 CAGCGAAATT CGTGGAAGAT GGmsAAAAGG TTAAGGCTGG CGACAAGCTA301 TTTGCGCTTT CGACCTCACG TTTCGGCGCA GGAGGTAGCG TGCAGCAGCA 351GTTGAAAACG GAGGCAGTTT TGAAGAAAAC GTTGGCAGAA CAGGAACTGG 401 GTCGTCTGAAGCTGATACAC GGGAATGAAA CGCGCAgCcT TAAAGCAACT 451 GTCGAACGTT TGGAAAACCAGGAACTCCAT ATTTCGCAAC AGATAGACGG 501 TCAGAAAAGG CGCATTAGAC TTGCGGAAGAAATGTTGCAG AAATATCGTT 551 TCCTATCCGC .CAATGA

This corresponds to the amino acid sequence <SEQ ID 420; ORF107>:

1 MNRPKQPFFR PEVAVARQTS LTGKVILTRP LSFSLWTTFA SISALLIILF 51 LIFGNYTRKTTVEGQILPAS GVIRVYAPDT XTITAKFVED GXKVKAGDKL 101 FALSTSRFGA GGSVQQQLKTEAVLKKTLAE QELGRLKLIH GNETRSLKAT 151 VERLENQELH ISQQIDGQKR RIRLAEEMLQKYRFLSXQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF107 shows 97.8% identity over a 186aa overlap with an ORF (ORF107a)from strain A of N. meningitidis:

The complete length ORF107a nucleotide sequence <SEQ ID 421> is:

1 ATGAATAGAC CCAAGCAACC NTTCTTCCGT CCCGAAGTCG CCGTTGCCCG 51 CCAAACCAGCCTGACGGGTA AAGTGATTCT GACACGACCG TTGTCATTTT 101 CCCTATGGAC GACATTTGCATCGATATCTG CGTTATTGAT TATCCTGTTT 151 TTGATATTTG GTAACTATAC GCGAAAGACAACAGTGGAGG GACAAATTTT 201 ACCTGCATCG GGCGTAATCA GGGTGTATGC ACCGGATACGGGGACAATTA 251 CNGCGAAATT CNTGGAAGAT GGAGAAAAGG TTAAGGCTGG CGACAAGCTA301 TTTGCGCTTT CGACCTCACG TTTCGGCGCA GGAGATAGCG TGCAGCAGCA 351GTTGAAAACG GAGGCAGTTT TGAAGAAAAC GTTGGCAGAA CAGGAACTGG 401 GTCGTCTGAAGCTGATACAC GGGAATGAAA CGCGCAGCCT TAAAGCAACT 451 GTCGAACGTT TGGAAAACCAGGAACTCCAT ATTTCGCAAC AGATAGACGG 501 TCAGAAAAGG CGCATTAGAC TTGCGGAAGAAATGTTGCAG AAATATCGTT 551 TCCTATCCGC CAATGATGCA GTGCCAAAAC AAGAAATGATGAATGTCAAG 601 GCAGAGCTTT TAGAGCAGAA AGCCAAACTT GATGCCTACC GCCGAGAAGA651 AGTCGGGCTG CTTCAGGAAA TCCGCACGCA GAATCTGACA TTGGNNAGCC 701TCCCCCAAGC GGCATGA

This encodes a protein having amino acid sequence <SEQ ID 422>:

1 MNRPKQPFFR PEVAVARQTS LTGKVILTRP LSFSLWTTFA SISALLIILF 51 LIFGNYTRKTTVEGQILPAS GVIRVYAPDT GTITAKFXED GEKVKAGDKL 101 FALSTSRFGA GDSVQQQLKTEAVLKKTLAE QELGRLKLIH GNETRSLKAT 151 VERLENQELH ISQQIDGQKR RIRLAEEMLQKYRFLSANDA VPKQEMMNVK 201 AELLEQKAKL DAYRREEVGL LQEIRTQNLT LXSLPQAA*Homology with a Predicted ORF from N. gonorrhoeae

ORF107 shows 95.7% identity over a 188aa overlap with a predicted ORF(ORF107.ng) from N. gonorrhoeae:

The complete length ORF107ng nucleotide sequence <SEQ ID 423> ispredicted to encode a protein having amino acid sequence <SEQ ID 424>:

1 MNRPKQPFFR PEVAIARQTS LTGKVILTRP LSFSLWTTFA SISALLIILF 51 LIFGNYTRKTTMEGQILPAS GVIRVYAPDT GTITAKFVED GEKVKAGDKL 101 FALSTSRFGA GGSVQQQLKTEAVLKKTLAE QELGRLKLIH ENETRSLKAT 151 VERLENQKLH ISQQIDGQKR RIRLAEEMLRKYRFLSAQ*

Based on the presence of a putative transmembrane domain in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 50

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 425>:

1 ATGCTGAATA CTTTTTTTGC CGTATTGGGC GGCTGCCTGC TGCT.TTGCC 51 GTGCGGCAAATCCGTAAATA CGGCGGTACA GCCGCAAAAC GCGGTACAAA 101 GCGCGCCGAA ACCGGTTTTCAAAGTCATAT ATATCGACAA TACGGCGATT 151 GCCGGTTTGG ATTTGGGACA AAGCAGCGAAGGCAAAACCA ACGACGGCAA 201 AAAACAAATC AGTTATCCGA TTAAAGGCTT GCCGGAACAAAATGTTATCC 251 GACTGATCGG CAAGCATCCC GGCGACTTGG AAGCCGTCAG CGGCAAATGT301 ATGGAAACCG ATGATAAGGA CAGTCCGGCA GGTTGGGCAG AAAACGGCGT 351GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC GAAGACGGCG 401 GCAAACTGACGGATTACCTA GTTTCGCATG CCGCCCTGCA ACCCTATCAG 451 GCAGGCAAAA GCGGCTATGCCGCCGTGCAG AACGGACGCT ATGTGCTGGA 501 AATCGACAGC GAAGGGGCGT TTTATTTCCGCCGCCGCCAT TATTGA

This corresponds to the amino acid sequence <SEQ ID 426; ORF108>:

1 MLNTFFAVLG GCLLXLPCGK SVNTAVQPQN AVQSAPKPVF KVIYIDNTAI 51 AGLDLGQSSEGKTNDGKKQI SYPIKGLPEQ NVIRLIGKHP GDLEAVSGKC 101 METDDKDSPA GWAENGVCHTLFAKLVGNIA EDGGKLTDYL VSHAALQPYQ 151 AGKSGYAAVQ NGRYVLEIDS EGAFYFRRRH Y*

Further work revealed the following DNA sequence <SEQ ID 427>:

1 ATGCTGAAAA CATCTTTTGC CGTATTGGGC GGCTGCCTGC TGCTTGCCGC 51 CTGCGGCAAATCCGAAAATA CGGCGGAACA GCCGCAAAAC GCGGTACAAA 101 GCGCGCCGAA ACCGGTTTTCAAAGTCAAAT ATATCGACAA TACGGCGATT 151 GCCGGTTTGG ATTTGGGACA AAGCAGCGAAGGCAAAACCA ACGACGGCAA 201 AAAACAAATC AGTTATCCGA TTAAAGGCTT GCCGGAACAAAATGTTATCC 251 GACTGATCGG CAAGCATCCC GGCGACTTGG AAGCCGTCAG CGGCAAATGT301 ATGGAAACCG ATGATAAGGA CAGTCCGGCA GGTTGGGCAG AAAACGGCGT 351GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC GAAGACGGCG 401 GCAAACTGACGGATTACCTA GTTTCGCATG CCGCCCTGCA ACCCTATCAG 451 GCAGGCAAAA GCGGCTATGCCGCCGTGCAG AACGGACGCT ATGTGCTGGA 501 AATCGACAGC GAAGGGGCGT TTTATTTCCGCCGCCGCCAT TATTGA

This corresponds to the amino acid sequence <SEQ ID 428; ORF108-1>:

1 MLKTSFAVLG GCLLLAACGK SENTAEQPQN AVQSAPKPVF KVKYIDNTAI 51 AGLDLGQSSEGKTNDGKKQI SYPIKGLPEQ NVIRLIGKHP GDLEAVSGKC 101 METDDKDSPA GWAENGVCHTLFAKLVGNIA EDGGKLTDYL VSHAALQPYQ 151 AGKSGYAAVQ NGRYVLEIDS EGAFYFRRRH Y*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. gonorrhoeae

ORF108 shows 88.4% identity over a 181 aa overlap with a predicted ORF(ORF 108.ng) from N. gonorrhoeae:

ORF108-1 shows 92.3% identity with ORF108ng over the same 181 aaoverlap:

The complete length ORF108ng nucleotide sequence <SEQ ID 429> is:

1 ATGCTGAAAa tacctTTTGC CGTGTtgggc ggCtgcctGC TGCTTGCCGC 51 CTGCGGCAAATCCGAAAATa cggcggaACA GCCGCAAAAT gcggCACAAA 101 GCGCGCCGAA ACCGGTTTTCAAAGTCAAAT ACATCGACAA TACGGCGATT 151 GCCGGTTTGG CTTTGGGACA AAGTAGCGAAGGCAAAACCA acgacgGCAA 201 AAAACAAATC AGTTATccgA TTAAAGGCTT GCCGGAACAAAacgccgtcc 251 gGCTGACCGG AAAGCATCCC AACGACTTGG AagccgtcgT CGGCAAATGT301 ATGGAAACCG ACGGAAAGGA CGCGCCTTCG GGCTGGGCGG AAAACGGCGT 351GTGCCATACC TTGTTTGCCA AACTGGTGGG CAATATCGCC GAAGACGGCG 401 GCAAACTGACTGATTACCTG ATTTCGCATT CCGCCCTGCA ACCCTATCAG 451 GCAGGCAAAA GCGGCTATGCCGCCGTGCAG AACGGACGCT ATGTGCTGGA 501 AATCGACAGC GagggGGCGT TTTATttccgccgccgccat tattgA

This encodes a protein having amino acid sequence <SEQ ID 430>:

1 MLKIPFAVLG GCLLLAACGK SENTAEQPQN AAQSAPKPVF KVKYIDNTAI 51AGLALGQSSE GKTNDGKKQI SYPIKGLPEQ NAVRLTGKHP NDLEAVVGKC 101 METDGKDAPSGWAENGVCHT LFAKLVGNIA EDGGKLTDYL ISHSALQPYQ 151 AGKSGYAAVQ NGRYVLEIDSEGAFYFRRRH Y*

Based on this analysis, including the presence of a predictedprokaryotic membrane lipoprotein lipid attachment site (underlined) anda putative ATP/GTP-binding site motif A (P-loop, double-underlined) inthe gonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 51

The following DNA sequence was identified in N. meningitidis <SEQ ID431>:

1 ATGGAAGATT TATATATAAT ACTCGCTTTG GGTTTGGTTG CGATGATTGC 51 CGgATTTATCGATgcgatTg cGggCGGGGG TGGTTTGATT ACGCTGCCCG 101 CACTCTTGTT GGCAGGTATTCCTCCCGTGT CGGCAATTGC CACCAACAAG 151 CTGCAAgCAG CCGCTGCTAC GTTTTCAGCTACGGTTTCTT TTGCACGCAA 201 AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCAGCATCGTTTG 251 TAGGCGGCGT GGcCGGTGCA TTATCGGTCA GCTTGGTTTC CAAAGATATT301 CTgCTgGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCAC TGTATTTTGT 351GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC AGAATGTCTT 401 TTTTTCTGTTcGGGCTGACG GTCGC.ACCG CTTTTGGGTT TTTACGACGG 451 TGTGTTCGGA CCGGGTGTCGGCTCGTTTTT TCTGATTGCC TTTATTGTTT 501 TGCTCGGCTG CAAgCTGTTG AACGCGATGTCTTACACCAA ATTGGCGAAC 551 GTTGCCTGCA ATCTTGGTTC GCTATCGGTA TTCCTGCTGCACGGTTCGAT 601 TATTTTCCCG ATTGCGGCAA CGaTGGCGGT CGGTGCGTTT GTCGGtGCGA651 ATTTAgGTGC GAGATTTGCC GTaCgctTCG GTTCGAAGCT GATTAA

This corresponds to the amino acid sequence <SEQ ID 432; ORF109>:

1 MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI PPVSAIATNK 51 LQAAAATFSATVSFARKGLI DWKKGLPIAA ASFVGGVAGA LSVSLVSKDI 101 LLAVVPVLLI FVALYFVFSPKLDGSKEGKA RMSFFLFGLT VXTAFGFLRR 151 CVRTGCRLVF SDCLYCFARL QAVERDVLHQIGERCLQSWF AIGIPAARFD 201 YFPDCGNDGG RCVCRCEFRC EICRTLRFEA D*

Further work revealed the following DNA sequence <SEQ ID 433>:

1 ATGGAAGATT TATATATAAT ACTCGCTTTG GGTTTGGTTG CGATGATTGC 51 CGGATTTATCGATGCGATTG CGGGCGGGGG TGGTTTGATT ACGCTGCCCG 101 CACTCTTGTT GGCAGGTATTCCTCCCGTGT CGGCAATTGC CACCAACAAG 151 CTGCAAGCAG CCGCTGCTAC GTTTTCAGCTACGGTTTCTT TTGCACGCAA 201 AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCAGCATCGTTTG 251 TAGGCGGCGT GGCCGGTGCA TTATCGGTCA GCTTGGTTTC CAAAGATATT301 CTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCAC TGTATTTTGT 351GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC AGAATGTCTT 401 TTTTTCTGTTCGGGCTGACG GTCGCACCGC TTTTGGGTTT TTACGACGGT 451 GTGTTCGGAC CGGGTGTCGGCTCGTTTTTT CTGATTGCCT TTATTGTTTT 501 GCTCGGCTGC AAGCTGTTGA ACGCGATGTCTTACACCAAA TTGGCGAACG 551 TTGCCTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCACGGTTCGATT 601 ATTTTCCCGA TTGCGGCAAC GATGGCGGTC GGTGCGTTTG TCGGTGCGAA651 TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG ATTAAGCCGC 701TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT GATAGACGAG 751 AGAAATCCGCTGTATCAGAT GATTGTTTCG ATGTTTTAA

This corresponds to the amino acid sequence <SEQ ID 434; ORF109-1>:

1 MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI PPVSAIATNK 51 LQAAAATFSATVSFARKGLI DWKKGLPIAA ASFVGGVAGA LSVSLVSKDI 101 LLAVVPVLLI FVALYFVFSPKLDGSKEGKA RMSFFLFGLT VAPLLGFYDG 151 VFGPGVGSFF LIAFIVLLGC KLLNAMSYTKLANVACNLGS LSVFLLHGSI 201 IFPIAATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISISMAVKLLIDE 251 RNPLYQMIVS MF*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF109 shows 95.9% identity over a 147aa overlap with an ORF (ORF109a)from strain A of N. meningitidis:

The complete length ORF109a nucleotide sequence <SEQ ID 435> is:

1 ATGGAAGATT TATACATAAT ACTCGCTTTG GGTTTGGTTG CGATGATTGC 51 CGGATTTATCGATGCGATTG CGGGTGGGGG TGGTTTGATT ACGCTGCCTG 101 CACTCTTGTT GGCAGGTATTCCTCCCGTGT CGGCAATTGC CACCAACAAG 151 CTGCAAGCAG CCGCTGCTAC GTTTTCGGCTACGGTTTCTT TTGCACGCAA 201 AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCGGCAGCATCGTTTG 251 CAGGCGGCGT GGTCGGTGCA TTATCGGTCA GCTTGGTTTC CAAAGATATT301 CTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCGC TGTATTTTGT 351GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC AGAATGTCTT 401 TTTTTCTGTTCGGTCTGACG GTTGCACCAC TTTTGGGTTT TTACGACGGT 451 GTGTTCGGAC CGGGTGTCGGCTCGTTTTTT CTGATTGCCT TTATTGTTTT 501 GCTCGGCTGC AAGCTGTTGA ACGCGATGTCTTACACCAAA TTGGCGAACG 551 TTGCCTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCACGGTTCGATT 601 ATTTTCCCGA TTGCGGCAAC GATGGCGGTC GGTGCGTTTG TCGGTGCGAA651 TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG ATTAAGCCGC 701TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT GATAGACGAG 751 AGAAATCCGCTGTATCAGAT GATTGTTTCG ATGTTTTAA

This encodes a protein having amino acid sequence <SEQ ID 436>:

1 MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI PPVSAIATNK 51 LQAAAATFSATVSFARKGLI DWKKGLPIAA ASFAGGVVGA LSVSLVSKDI 101 LLAVVPVLLI FVALYFVFSPKLDGSKEGKA RMSFFLFGLT VAPLLGFYDG 151 VFGPGVGSFF LIAFIVLLGC KLNAMSYTKLANVACNLGS LSVFLLHGSI 201 IFPIAATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISISMAVKLLIDE 251 RNPLYQMIVS MF*

ORF109a and ORF109-1 show 99.2% identity in 262 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF109 shows 98.3% identity over a 231aa overlap with a predicted ORF(ORF109.ng) from N. gonorrhoeae:

An ORF109ng nucleotide sequence <SEQ ID 437> was predicted to encode aprotein having amino acid sequence <SEQ ID 438>:

1 MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI PPVSAIATNK 51 LQAAAATFSATVSFARKGLI DWKKGLPIAA ASFAGGVVGA LSVSLVSKDI 101 LLAVVPVLLI FVALYFVFSPKLDGSKEGKA RMSFFLFGLT VATAFGFLRR 151 CVRTGCRLVF SDCLYCFARL QAVERDVLHQIGERCLQSWF AIGIPAARFD 201 YFPDCGNDGG RCVCRCEFRC EICRPLRFEA D*

Further work revealed the following gonococcal DNA sequence <SEQ ID439>:

1 ATGGAAGATT TATACATAAT ACTCGCTTTG GGTTTGGTTG CGATGATCGC 51 CGGATTTATCGATGCGATTG CGGGCGGGGG TGGTTTGATT ACGCTGCCTG 101 CACTCTTGTT GGCAGGTATTCCTCCCGTGT CGGCAATTGC CACCAACAAG 151 CTGCAAGCAG CCGCTGCTAC GTTTTCGGCTACGGTTTCTT TTGCACGCAA 201 AGGTTTGATT GATTGGAAGA AAGGTCTCCC GATTGCCGCAGCATCGTTTG 251 CAGGCGGCGT GGTCGGTGCA TTATCGGTCA GCTTGGTTTC CAAAGATATT301 TTGCTGGCGG TCGTGCCGGT TTTGTTGATA TTTGTCGCGC TGTATTTTGT 351GTTTTCGCCC AAGCTCGACG GCAGTAAGGA AGGCAAAGCC AGAATGTCTT 401 TTTTTCTATTCGGGCTGACG GTTGCACCGC TTTTGGGTTT TTACGACGGT 451 GTGTTCGGAC CGGGTGTCGGCTCGTTTTTT CTGATTGCCT TTATTGTTTT 501 GCTCGGCTGC AAGCTGTTGA ACGCGATGTCTTACACCAAA TTGGCGAACG 551 TTGCTTGCAA TCTTGGTTCG CTATCGGTAT TCCTGCTGCACGGTTCGATT 601 ATTTTCCCGA TTGTGGCAAC GATGGCGGTC GGTGCGTTTG TCGGTGCGAA651 TTTAGGTGCG AGATTTGCCG TCCGCTTCGG TTCGAAGCTG ATTAAGCCGC 701TGCTGATTGT CATCAGCATT TCGATGGCTG TGAAATTGTT GATAGACGAG 751 AGAAATCCGCTGTATCAGAT GATTGTTTCG ATGTTTTAA

This corresponds to the amino acid sequence <SEQ ID 440; ORF109ng-1>:

1 MEDLYIILAL GLVAMIAGFI DAIAGGGGLI TLPALLLAGI PPVSAIATNK 51 LQAAAATFSATVSFARKGLI DWKKGLPIAA ASFAGGVVGA LSVSLVSKDI 101 LLAVVPVLLI FVALYFVFSPKLDGSKEGKA RMSFFLFGLT VAPLLGFYDG 151 VFGPGVGSFF LIAFIVLLGC KLLNAMSYTKLANVACNLGS LSVFLLHGSI 201 IFPIVATMAV GAFVGANLGA RFAVRFGSKL IKPLLIVISISMAVKLLIDE 251 RNPLYQMIVS MF*

ORF109ng-1 and ORF109-1 show 98.9% identity in 262 aa overlap:

In addition, ORF109ng-1 shows homology to a hypothetical Pseudomonasprotein:

sp|P29942|YCB9_PSEDE HYPOTHETICAL 27.4 KD PROTEIN IN COBO 3′REGION(ORF9) >gi|94984|pir||I38164 hypothetical protein 9 - Pseudomonassp >gi|551929 (M62866) ORF9 [Pseudomonas denitrificans] Length = 261Score = 175 bits (439), Expect = 3e−43 Identities = 83/214 (38%),Positives = 131/214 (60%), Gaps = 1/214 (0%) Query: 41PPVSAIATNKLQXXXXXXXXXXXXXRKGLIDWKKGLPIXXXXXXXXXXXXXXXXXXXKDI 100PP+  + TNKLQ             R+G ++ K+ LP+                    D+ Sbjct: 43PPLQTLGTNKLQGLFGSGSATLSYARRGHVNLKEQLPMALMSAAGAVLGALLATIVPGDV 102 Query:101 LLAVVPVLLIFVALYFVFSPKLDGSKEGKARMSFFLFGLTVAPLLGFYDGVFGPGVGSFF 160 LA++P LLI +ALYF   P + G  +  +R++ F+F LT+ PL+GFYDGVFGPG GSFF Sbjct: 103LKAILPFLLIAIALYFGLKPNM-GDVDQHSRVTPFVFTLTLVPLIGFYDGVFGPGTGSFF 161 Query:161 LIAFIVLLGCKLLNAMSYTKLANVACNLGSLSVFLLHGSIIFPIVATMAVGAFVGANLGA 220++ F+ L G  +L A ++TK  N   N+G+  VFL  G++++ +   M +G F+GA +G+ Sbjct: 162MLGFVTLAGFGVLKATAHTKFLNFGSNVGAFGVFLFFGAVLWKVGLLMGLGQFLGAQVGS 221 Query:221 RFAVRFGSKLIKPLLIVISISMAVKLLIDERNPL 254 R+A+  G+K+IKPLL+++SI++A++LLD  +PL Sbjct: 222 RYAMAKGAKIIKPLLVIVSIALAIRLLADPTHPL 255

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 52

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 441>:

1 ..CTGCTAGGGT ATTGCATCGG TTATCGGTAC GGCTGTTGCA GCAAAACCAG 51  CCGCAGACGG ATTATTTGGT CAAATTCGGA TCGTTTTGGG CGAG.ATTTT 101  TGGTTTTCTG GGACTGTATG ACGTCTATGC TTCGGCATGG TTTGTCGTTA 151  TCATGATGTT TTTGGTGGTT TCTACCAGTT TGTGCCTGAT TCGCAATGTG 201  CCGCCGTTCT GGCGCGAAAT GAAGTCTTTT CGGGAAAAGG TTAAAGAAAA 251  ATCTCTGGCG GCGATGCGCC ATTCTTCGCT GTTGGATGTA AAAATTGCGC 301  CCGAGGTTGC CAAACGTTAT CTGGAAGTAC AAGGTTTTCA GGGGAAAACC 351  ATTAACCGTG AAGACGGGTC GGTTCTGATT GCCGCCAAAA AAGGCACAAT 401  GAACAAATGG GGCTATATCT TTGCCCATGT TGCTTTGATT GTCATTTGCC 451  TGGGCGGGTT GATAGACAGT AACCTGCTGT TGAAACTGGG TATGCTGACC 501  GGTCGGATTG TTCCGGACAA TCAGGCGGTT TATGCCAAGG ATTTC.AAGC 551  CCGAAAGTAT .TTTGGGTGC gTCCAATCTC TCATTTAGGG GCAACGTCAA 601  TATTTCCG.A GGGGCAGAgT GCGGATGTGG TTTTCCTGA

This corresponds to the amino acid sequence <SEQ ID 442; ORF110>:

1 ..LLGIASVIGT LLQQNQPQTD YLVKFGSFWA XIFGFLGLYD VYASAWFVVI 51  MMFLVVSTSL CLIRNVPPFW REMKSFREKV KEKSLAAMRH SSLLDVKIAP 101  EVAKRYLEVQ GFQGKTINRE DGSVLIAAKK GTMNKWGYIF AHVALIVICL 151  GGLIDSNLLL KLGMLTGRIF RTIRRFMPRI XKPESXFGCV QSLI*GQRQY 201  FXRGRVRMWF S*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with ORF88a from N. meningitidis (Strain A)

ORF110 shows 91.5% identity over a 188aa overlap with ORF88a from strainA of N. meningitidis:

However, ORF88 and ORF110 do not align, because they represent twodifferent fragments of the same protein.

Homology with a Predicted ORF from N. gonorrhoeae

ORF110 shows 88.6% identity over a 211 as overlap with a predicted ORF(ORF110.ng) from N. gonorrhoeae:

The complete length ORF110ng nucleotide sequence <SEQ ID 443> ispredicted to encode a protein having amino acid sequence <SEQ ID 444>:

1 MSKSRISPTL LSRPWFAFFS SMRFAVALLS LLGIASVIGT VLQQNQPQTD 51 YLVKFGPFWTRIFDFLGLYD VYASAWFVVI MMFLVVSTSL CLIRNVPPFW 101 REMKSFREKV KEKSLAAMRHSSLLDVKIAP EVAKRYLEVR GFQGKTVSRE 151 DGSVLIAAKK GTMNKWGYIXAHVALIVICL GRLINXNLLL KLGMLAGSIF 201 RNNRRVMPRI SKPESIWGGV QSLIKGQRQYFQRGKVRMWF S*

Based on the putative transmembrane domains in the gonococcal protein,it is predicted that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 53

The following DNA sequence was identified in N. meningitidis <SEQ ID445>:

1 ATGCCGTCTG AAACACGCCT GCCGAACTTT ATCCGCGTCT TGATATTTGC 51 CCTGGGTTTCATCTTCCTGA ACGCCTGTTC GGAACAAACC GCGCAAACCG 101 TTACCCTGCA AGGCGAAACGATGGGCACGA CCTATACCGT CAAATACCTT 151 TCAAATAATC GGGACAAACT CCCCTCACCTGCCGAAATAC AAAAACGCAT 201 CGATGACGCG CTTAAAGAAG TCAACCGGCA GATGTCCACCTATCAGCCCG 251 ACTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA GCCCCTCCGC301 ATTTCAAGCG ACTTCGCACA CGTTACTGCC GAAGCCGTCC GCCTGAACCG 351CCTGACACAC GGCGCGCTGG ACGTAACCGT CGGCCCCTTG GTCAACCTTT 401 GGGGATTCGGCCCCGACAAA TCCGTTACCC GTGAACCGTC GCCGGAACAA 451 ATCAAACAGG CGGCATCTTATACGGGCATA GACAAAATCA TTTTGAAACA 501 AGGCAAAGAT TACGCTTCCT TGAGCAAAACCCACCCCAAG GCCTATTTGG 551 ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATAAAGTTGCGGGCGAA 601 CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAATCG GCGGCGAGTT651 GCACGGCAAA GGCAAAAACG CGCGCGGCGA ACCGTGGCGC ATCGGTATCG 701AGCAGCCCAA TATCGTCCAA GGCGGCAATA CGCAGATTAT CGTCCCGCTG 751 AACAACCGTTCGCTTGCCAC TTCCGGCGAT TACCGTATTT TCCACGTCGA 801 TAAAAACGGC AAACGCCTCTCCCATATCAT CAACCCGAAC AACAAACGAC 851 CCATCAGCCA CAACCTCGCC TCCATCAGCGTGGTCGCAGA CAGTGCGATG 901 ACGGCGGACG GCTTGTCCAC AGGATTATTC GTATTGGGCGAAACCGAAGC 951 CTTAAAGCTG GCAGAGCGCG AAAAACTCGC TGTTTTCCTG ATTGTCAGGG1001 ATAAAGGCGG CTACCGCACC GCCATGTCTT CCGAATTTGA AAAACTGCTC 1051 CGCTAA

This corresponds to the amino acid sequence <SEQ ID 446; ORF111>:

1 MPSETRLPNF IRVLIFALGF IFLNACSEQT AQTVTLQGET MGTTYTVKYL 51 SNNRDKLPSPAEIQKRIDDA LKEVNRQMST YQPDSEISRF NQHTAGKPLR 101 ISSDFAHVTA EAVRLNRLTHGALDVTVGPL VNLWGFGPDK SVTREPSPEQ 151 IKQAASYTGI DKIILKQGKD YASLSKTHPKAYLDLSSIAK GFGVDKVAGE 201 LEKYGIQNYL VEIGGELHGK GKNARGEPWR IGIEQPNIVQGGNTQIIVPL 251 NNRSLATSGD YRIFHVDKNG KRLSHIINPN NKRPISHNLA SISVVADSAM301 TADGLSTGLF VLGETEALKL AEREKLAVFL IVRDKGGYRT AMSSEFEKLL 351 R*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF111 shows 96.9% identity over a 351aa overlap with an ORF (ORF111a)from strain A of N. meningitidis:

The complete length ORF111a nucleotide sequence <SEQ ID 447> is:

1 ATGCCGTCTG AAACACGCCT GCCGAACTTT ATCCGCACCT TGATATTTGC 51 CCTGAGTTTTATCTTCCTGA ACGCCTGTTC GGAACAAACC GCGCAAACCG 101 TTACCCTGCA AGGTGAAACGATGGGCACGA CCTATACCGT CAAATACCTT 151 TCAAATAATC GGGACNAACT CCCNTCACCTGCCGAAATAC AAAANCGCAT 201 CGATGACGCG CTTAAAGAAG TCAACCGGCA GATGTCCACCTATCAGCCCG 251 ACTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA GCCCCTCCGC301 ATTTCAAGCG ACTTCGCACA CGTTACTGCC GAAGCCGTCC ACCTGAACCG 351CCTGACACAC GGCGCGCTGG ACGTAACCGT CGGCCCCTTG GTCAACCTTT 401 GGGGATTCGGCCCCGACAAA TCCGTTACCC GTGAACCGTC GCCGGAACAA 451 ATCAAACAAG CAGCATCTTATACGGGCATA GACAAAATCA TTTTGAAACA 501 AGGCAAAGAT TACGCTTCCT TGAGCAAAACCCACCCCAAG GCCTATTTGG 551 ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATNANGTTGCGGGCGAA 601 CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAATCG GCGGNGAGTT651 GCACGGCAAA GNCAAAAACG CGCGCGGCGA ACCTTGGCGC ATCGGCATCG 701AACAGCCCAA CATCGTCCAA GGCGGCAATA CGCAGATTAT CGTCCCGCTG 751 AACAACCGTTCGNTTGCCAC TTCCGGCGAT TACCGTATTT TCCACGTCGA 801 TAAAAGCGGC AAACGCCTCTCCCATATCAT TAATCCGAAC AACAAACGAC 851 CCATCAGCCA CAACCTCGCC TCCATCAGCGTGNTCGCAGA CAGTGCGATG 901 ACGGCGGACG GCTTNTCCAC AGGATTATTC GTATTGGGCGAAACCGAAGC 951 CTTAAAGCTG GCAGAGCGCG AAAAACTCGC TGTTTTCCTG ATTGTCAGGG1001 ATAAAGGCGG CTACCGCACC GCCATGTCTT CCGAATTTGA AAAACTGCTC 1051 CGCTAA

This encodes a protein having amino acid sequence <SEQ ID 448>:

1 MPSETRLPNF IRTLIFALSF IFLNACSEQT AQTVTLQGET MGTTYTVKYL 51 SNNRDXLPSPAEIQXRIDDA LKEVNRQMST YQPDSEISRF NQHTAGKPLR 101 ISSDFAHVTA EAVHLNRLTHGALDVTVGPL VNLWGFGPDK SVTREPSPEQ 151 IKQAASYTGI DKIILKQGKD YASLSKTHPKAYLDLSSIAK GFGVDXVAGE 201 LEKYGIQNYL VEIGGELHGK XKNARGEPWR IGIEQPNIVQGGNTQIIVPL 251 NNRSXATSGD YRIFHVDKSG KRLSHIINPN NKRPISHNLA SISVXADSAM301 TADGXSTGLF VLGETEALKL AEREKLAVFL IVRDKGGYRT AMSSEFEKLL 351 R*Homology with a Predicted ORF from N. gonorrhoeae

ORF111 shows 96.6% identity over a 351aa overlap with a predicted ORF(ORF111.ng) from N. gonorrhoeae:

The complete length ORF111ng nucleotide sequence <SEQ ID 449> is:

1 ATGCCGTCTG AAACACGCCT GCCGAACCTT ATCCGCGCCT TGATATTTGC 51 CCTGGGTTTCATCTTCCTGA ACGCCTGTTC GGaacaaacC GCGCAaaccg 101 TTACCCTGCA AGGCGAAAcgaTGGGTACGA CCTATACCGT CAAATACCTT 151 TCAAATAATC GGGACAAACT CCCCTCCCCTGCCAAAATAC AAAAGCGCAT 201 TGATGATGCG CTTAAAGAAG TCAACCGGCA GATGTCCACCTACCAGACCG 251 ATTCCGAAAT CAGCCGGTTC AACCAACACA CAGCCGGCAA GCCCCTCCGC301 ATTTCAAGCG ATTTCGCACA CGTTACCGCC GAAGCCGTCC GCCTGAACCG 351CCTGACTCAC GGCGCACTGG ACGTAACCGT CGGCCCTTTG GTCAACCTTT 401 GGGGGTTCGGCCCCGACAAA TCCGTTACCC GTGAACCGTC GCCGGAACAA 451 ATCAAACAGG CGGCATCTTATACGGGCATA GACAAAATCA TTTTGCAACA 501 AGGCAAAGAT TACGCTTCCT TGAGCAAAACCCACCCCAAA GCCTATTTGG 551 ATTTATCTTC GATTGCCAAA GGCTTCGGCG TTGATAAAGTTGCGGGCGAA 601 CTGGAAAAAT ACGGCATTCA AAATTATCTG GTCGAAAtcg gcggcGAGTT651 GCACGGCAAA GGCAAAAATG CGCACGGCGA ACCGTGGCGC ATCGGTATAG 701AGCAACCCAA TATCATCCAA GgcgGCAata CGCAGATTAt cgtcccgctg 751 aaCaaccgttcgctTGCCAC TTCCGGCGAT TAccgtaTTT tccacgtcgA 801 TAAAAAcggc aaacgcctttcccacaTCAT CAATCCCaAC aacAAACgac 851 ccATCAGcca caacctcgcc tccatcagcgtggtctcAGA CAGTGCAATG 901 ACGGCGGACG GTTtatCCAC AGGATTATTT GTTTTAGGCGAAACCGAAGC 951 CTTAAGGCTG GCAGAACAAG AAAAACTCGC TGTTTTCCTA ATTGTCCGGG1001 ATAAGGACGG CTACCGCACC GCCATGTCTT CCGAATTTGC CAAGCTGCTC 1051 CGCTAA

This encodes a protein having amino acid sequence <SEQ ID 450>:

1 MPSETRLPNL IRALIFALGF IFLNACSEQT AQTVTLQGET MGTTYTVKYL 51 SNNRDKLPSPAKIQKRIDDA LKEVNRQMST YQTDSEISRF NQHTAGKPLR 101 ISSDFAHVTA EAVRLNRLTHGALDVTVGPL VNLWGFGPDK SVTREPSPEQ 151 IKQAASYTGI DKIILQQGKD YASLSKTHPKAYLDLSSIAK GFGVDKVAGE 201 LEKYGIQNYL VEIGGELHGK GKNAHGEPWR IGIEQPNIIQGGNTQIIVPL 251 NNRSLATSGD YRIFHVDKNG KRLSHIINPN NKRPISHNLA SISVVSDSAM301 TADGLSTGLF VLGETEALRL AEQEKLAVFL IVRDKDGYRT AMSSEFAKLL 351 R*

This protein shows homology with a hypothetical lipoprotein precursorfrom H. influenzae:

sp|P44550|YOJL_HAEIN HYPOTHETICAL LIPOPROTEIN HI0172PRECURSOR >gi|1074292|pir|4 hypothetical protein HI0172 - Haemophilusinfluenzae (strain Rd KW20) >gi|1573128 (U32702) hypothetical[Haemophilus influenzae] Length = 346 Score = 353 bits (896), Expect= 9e−97 Identities = 181/344 (52%), Positives = 247/344 (71%), Gaps= 4/344 (1%) Query: 7LPNLIRALIFALGFIFLNACSEQTAQTVTLQGETMGTTYTVKYLSNNRDKLPSPAKIQKR 66+  LI  +I     + L AC ++T + ++L G+TMGTTY VKYL +      S  K  + Sbjct: 1MKKLISGIIAVAMALSLAACQKET-KVISLSGKTMGTTYHVKYLDDGSITATSE-KTHEE 58 Query:67 IDDALKEVNRQMSTYQTDSEISRFNQHT-AGKPLRISSDFAHVTAEAVRLNRLTHGALDV 125I+  LK+VN +MSTY+ DSE+SRFNQ+T    P+ IS+DFA V AEA+RLN++T GALDV Sbjct: 59IEAILKDVNAKMSTYKKDSELSRFNQNTQVNTPIEISADFAKVLAEAIRLNKVTEGALDV 118 Query:126 TVGPLVNLWGFGPDKSVTREPSPEQIKQAASYTGIDKIILQQGKDYASLSKTHPKAYLDL 185TVGP+VNLWGFGP+K   ++P+PEQ+ +  ++ GIDKI L   K+ A+LSK  P+ Y+DL Sbjct: 119TVGPVVNLWGFGPEKRPEKQPTPEQLAERQAWVGIDKITLDTNKEKATLSKALPQVYVDL 178 Query:186 SSIAKGFGVDKVAGELEKYGIQNYLVEIGGELHGKGKNAHGEPWRIGIEQPNIIQGGNTQ 245SSIAKGFGVD+VA +LE+   QNY+VEIGGE+  KGKN  G+PW+I IE+P        + Sbjct: 179SSIAKGFGVDQVAEKLEQLNAQNYMVEIGGEIRAKGKNIEGKPWQIAIEKPTTTGERAVE 238 Query:246 IIVPLNNRSLATSGDYRIFHVDKNGKRLSHIINPNNKRPISHNLASISVVSDSAMTADGL 305 ++ LNN  +A+SGDYRI+  ++NGKR +H I+P    PI H+LASI+V++ ++MTADGL Sbjct: 239AVIGLNNMGMASSGDYRIY-FEENGKRFAHEIDPKTGYPIQHHLASITVLAPTSMTADGL 297 Query:306 STGLFVLGETEALRLAEQEKLAVFLIVRDKDGYRTAMSSEFAKL 349 STGLFVLGE +AL+AE+  LAV+LI+R  +G+ T  SS F KL Sbjct: 298STGLFVLGEDKALEVAEKNNLAVYLIIRTDNGFVTKSSSAFKKL 341

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 54

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 451>:

1 ..CCGTGCCGCC GACAGGGCGA CGACGTGTAT GCGGCGCACG CGTCCCGTCA 51  AAAATTGTGG CTGCGCTTCA TCGGCGGCCG GTCGCATCAA AATATACGGG 101  GCGGCGCGGC TGCGGACGGG TGGCGCAAAG GCGTGCAAAT CGGCGGCGAG 151  GTGTTTGTAC GGCAAAATGA AGGCAGCCkA yTGGCAATCG GCGTGATGGG 201  CGGCAGGGCC GGCCAGCACG CwTCAGTCAA CGGCAAAGGC GGTGCGGCAG 251  gCAGTGATTT GTATGGTTAT GgCGGGGgTG TTTATGCTgC GTGGCATCAG 301  TTGCGCGATA AACAAACGGG TgCGTATTTG GACGGCTGGT TGCAATACCA 351  ACGTTTCAAA CACCGCATCA ATGATGAAAA CCGTGCGGAA CgCTACAAAA 401  CCAAAGGTTG GACGGCTTCT GTCGAAGGCG GCTACAACGC GCTTGTGGCG 451  GAAGGCATTG TCGGAAAAGG CAATAATGTG CGGTTTTACC TACAACCGCA 501  GgCGCAGTTT ACCTACTTGG GCGTAAACGG CGGCTTTACC GACAGCGAGG 551  GGACGGCGGT CGGACTGCTC GGCAGCGGTC AGTGGCAAAG CCGCGCCGGC 601  AtTCGGGCAA AAACCCGTTT TGCTTTGCGT AACGGTGTCA ATCTTCAGCC 651  TTTTGCCGCT TTTAATGTtt TGCACAGGTC AAAATCTTTC GGCGTGGAAA 701  TGGACGGCGA AAAACAGACG CTGGCAGGCA GGACGGCACT CGAAGGGCGG 751  TTCGGTATTG AAGCCGGTTG GAAAGGCCAT ATGTCCGCA..

This corresponds to the amino acid sequence <SEQ ID 452; ORF35>:

1 ..PCRRQGDDVY AAHASRQKLW LRFIGGRSHQ NIRGGAAADG WRKGVQIGGE 51  VFVRQNEGSX LAIGVMGGRA GQHASVNGKG GAAGSDLYGY GGGVYAAWHQ 101  LRDKQTGAYL DGWLQYQRFK HRINDENRAE RYKTKGWTAS VEGGYNALVA 151  EGIVGKGNNV RFYLQPQAQF TYLGVNGGFT DSEGTAVGLL GSGQWQSRAG 201  IRAKTRFALR NGVNLQPFAA FNVLHRSKSF GVEMDGEKQT LAGRTALEGR 251  FGIEAGWKGH MSA..

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Putative Secreted VirG-Homolgue of N. meningitidis(Accession Number A32247)

ORF and virg-h protein show 51% aa identity in 261aa overlap:

Orf35 5 QGDDVYAAHASRQKLWLRFIGGRSHQNIRGGAA-ADGWRKGVQIGGEVFVRQNEGSXLAI 63+  D++     R+ LWLR I G S+Q ++G  A  +G+RKGVQ+GGEVF  QNE + L+I virg-h 396KNSDIFDRTLPRKGLWLRVIDGHSNQWVQGKTAPVEGYRKGVQLGGEVFTWQNESNQLSI 455 Orf3564 GVMGGRAGQHASVNGKG--GAAGSDLYGYGGGVYAAWHQLRDKQTGAYLDGWLQYQRFKH 121G+MGG+A Q ++ +          ++ G+G GVYA WHQL+DKQTGAY D W+QYQRF+H virg-h 456GLMGGQAEQRSTFHNPDTDNLTTGNVKGFGAGVYATWHQLQDKQTGAYADSWMQYQRFRH 515 Orf35122 RINDENRAERYKTKGWTASVEGGYNALVAEGIVGKGNNVRFYLQPQAQFTYLGVNGGFTD 181 RINE+  ER+ +KG TAS+E GYNAL+AE    KGN++R YLQPQAQ TYLGVNG F+D virg-h 516RINTEDGTERFTSKGITASIEAGYNALLAEHFTKKGNSLRVYLQPQAQLTYLGVNGKFSD 575 Orf35182 SEGTAVGLLGSGQWQSRAGIRAKTRFALRNGVNLQPFAAFNVLHRSKSFGVEMDGEKQTL 241SE   V LLGS Q Q+R G++AK +F+L   + ++PFAA N L+ +K FGVEMDGE++ + virg-h 576SENAHVNLLGSRQLQTRVGVQAKAQFSLYKNIAIEPFAAVNALYHNKPFGVEMDGERRVI 635 Orf35242 AGRTALEGRFGIEAGWKGHMS 262   +TA+E + G+    K H++ virg-h 636NNKTAIESQLGVAVKIKSHLT 656Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF35 shows 96.9% identity over a 259aa overlap with an ORF (ORF35a)from strain A of N. meningitidis:

The complete length ORF35a nucleotide sequence <SEQ ID 453> is:

1 ATGTTCAGAG CTCAGCTTGG TTCAAATACT CGTTCTACCA AAATCGGCGA 51 CGATGCCGATTTTTCATTTT CAGACAAGCC GAAACCCGGC ACTTCCCATT 101 ATTTTTCCAG CGGTAAAACCGATCAAAATT CATCCGAATA TGGGTATGAC 151 GAAATCAATA TCCAAGGTAA AAACTACAATAGCGGCATAC TCGCCGTCGA 201 TAATATGCCC GTTGTTAAGA AATATATTAC AGATACTTACGGGGATAATT 251 TAAAGGATGC GGTTAAGAAG CAATTACAGG ATTTATACAA AACAAGACCC301 GAAGCTTGGG AAGAAAATAA AAAACGGACT GAGGAGGCGT ATATAGAACA 351GCTTGGACCA AAATTTAGTA TACTCAAACA GAAAAACCCC GATTTAATTA 401 ATAAATTGGTAGAAGATTCC GTACTCACTC CTCATAGTAA TACATCACAG 451 ACTAGTCTCA ACAACATCTTCAATAAAAAA TTACACGTCA AAATCGAAAA 501 CAAATCCCAC GTCGCCGGAC AGGTGTTGGAACTGACCAAG ATGACGCTGA 551 AAGATTCCCT TTGGGAACCG CGCCGCCATT CCGACATCCATATGCTGGAA 601 ACTTCCGATA ATGCCCGCAT CCGCCTGAAC ACGAAAGATG AAAAACTGAC651 CGTCCATAAA GCGTATCAGG GCGGTGCGGA TTTCCTGTTC GGCTACGACG 701TGCGGGAGTC GGACAAACCC GCCCTGACCT TTGAAGAAAA AGTCAGCGGA 751 CAATCCGGCGTGGTTTTGGA ACGCCGGCCG GAAAATCTGA AAACGCTCGA 801 CGGGCGCAAA CTGATTGCGGCGGAAAAGGC AGACTCTAAT TCGTTTGCGT 851 TTAAACAAAA TTACCGGCAG GGACTGTACGAATTATTGCT CAAGCAATGC 901 GAAGGCGGAT TTTGCTTGGG CGTGCAGCGT TTGGCTATCCCCGAGGCGGA 951 AGCGGTTTTA TATGCCCAAC AGGCTTATGC GGCAAATACT TTGTTCGGGC1001 TGCGTGCCGC CGACAGGGGC GACGACGTGT ATGCCGCCGA TCCGTCCCGT 1051CAAAAATTGT GGCTGCGCTT CATCGGCGGC CGGTCGCATC AAAATATACG 1101 GGGCGGCGCGGCTGCGGACG GGCGGCGCAA AGGCGTGCAA ATCGGCGGCG 1151 AGGTGTTTGT ACGGCAAAATGAAGGCAGCC GGCTGGCAAT CGGCGTGATG 1201 GGCGGCAGGG CTGGCCAGCA CGCATCAGTCAACGGCAAAG GCGGTGCGGC 1251 AGGCAGTTAT TTGCATGGTT ATGGCGGGGG TGTTTATGCTGCGTGGCATC 1301 AGTTGCGCGA TAAACAAACG GGTGCGTATT TGGACGGCTG GTTGCAATAC1351 CAACGTTTCA AACACCGCAT CAATGATGAA AACCGTGCGG AACGCTACAA 1401AACCAAAGGT TGGACGGCTT CTGTCGAAGG CGGCTACAAC GCGCTTGTGG 1451 CGGAAGGCGTTGTCGGAAAA GGCAATAATG TGCGGTTTTA CCTGCAACCG 1501 CAGGCGCAGT TTACCTACTTGGGCGTAAAC GGCGGCTTTA CCGACAGCGA 1551 GGGGACGGCG GTCGGACTGC TCGGCAGCGGTCAGTGGCAA AGCCGCGCCG 1601 GCATTCGGGC AAAAACCCGT TTTGCTTTGC GTAACGGTGTCAATCTTCAG 1651 CCTTTTGCCG CTTTTAATGT TTTGCACAGG TCAAAATCTT TCGGCGTGGA1701 AATGGACGGC GAAAAACAGA CGCTGGCAGG CAGGACGGCG CTCGAAGGGC 1751GGTTCGGCAT TGAAGCCGGT TGGAAAGGCC ATATGTCCGC ACGCATCGGA 1801 TACGGCAAAAGGACGGACGG CGACAAAGAA GCCGCATTGT CGCTCAAATG 1851 GCTGTTTTGA

This encodes a protein having amino acid sequence <SEQ ID 454>:

1 MFRAQLGSNT RSTKIGDDAD FSFSDKPKPG TSHYFSSGKT DQNSSEYGYD 51 EINIQGKNYNSGILAVDNMP VVKKYITDTY GDNLKDAVKK QLQDLYKTRP 101 EAWEENKKRT EEAYIEQLGPKFSILKQKNP DLINKLVEDS VLTPHSNTSQ 151 TSLNNIFNKK LHVKIENKSH VAGQVLELTKMTLKDSLWEP RRHSDIHMLE 201 TSDNARIRLN TKDEKLTVHK AYQGGADFLF GYDVRESDKPALTFEEKVSG 251 QSGVVLERRP ENLKTLDGRK LIAAEKADSN SFAFKQNYRQ GLYELLLKQC301 EGGFCLGVQR LAIPEAEAVL YAQQAYAANT LFGLRAADRG DDVYAADPSR 351QKLWLRFIGG RSHQNIRGGA AADGRRKGVQ IGGEVFVRQN EGSRLAIGVM 401 GGRAGQHASVNGKGGAAGSY LHGYGGGVYA AWHQLRDKQT GAYLDGWLQY 451 QRFKHRINDE NRAERYKTKGWTASVEGGYN ALVAEGVVGK GNNVRFYLQP 501 QAQFTYLGVN GGFTDSEGTA VGLLGSGQWQSRAGIRAKTR FALRNGVNLQ 551 PFAAFNVLHR SKSFGVEMDG EKQTLAGRTA LEGRFGIEAGWKGHMSARIG 601 YGKRTDGDKE AALSLKWLF*Homology with a Predicted ORF from N. gonorrhoeae

ORF35 shows 51.7% identity over a 261aa overlap with a predicted ORF(ORF35ngh) from N. gonorrhoeae:

A partial ORF35ngh nucleotide sequence <SEQ ID 455> is predicted toencode a protein having partial amino acid sequence <SEQ ID 456>:

1 ..KKLRDRNSEY WKEETYHIKS NGRTYPNIPA LFPKHPFDPF ENINNSKKIS 51  FYDKEYTEDY LVGFARGFGV EKRNGEEEKP LRQYFKDCVN TENSNNDNCK 101  ISSFGNYGPI LIKSDIFALA SQIKNSHINS EILSVGNYIE WLRPTLNKLT 151  GWQEHLYAGL DPFHYIEVTD NSHVIGQTID LGALELTNSL WKPRWNSNID 201  YLITKNAEIR FNTKNESLLV KEDYAGGARF RFAYDLKDKV PEIPVLTFEK 251  NITGTSDIIF EGKALDNLKH LDGHQIVKVN DTADKDAFRL SSKYRKGIYT 301  LSLQQRPEGF FTKVQERDDI AIYAQQAQAA NTLFALRLND KNSDIFDRTL 351  PRKGLWLRVI DGHSNQWVQG KTAPVEGYRK GVQLGGEVFT WQNESNQLSI 401  GLMGGQAEQR STFRNPDTDN LTTGNVKGFG AGVYATWHQL QDKQTGAYVD 451  SWMQYQRFRH RINTEYATER FTSKGITASI EAGYNALLAE HFTKKGNSLR 501  VYLQPQAQLT YLGVNGKFSD SENAQVNLLG SRQLQSRVGV QAKAQFAFTN 551  GVTFQPFVAV NSIYQQKPFG VEIDGDRRVI NNKTVIETQL GVAAKIKSHL 601  TLQASFNRQT SKHHHAKQGA LNLQWTF*

Based on this prediction, these proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 55

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 457>:

  1 ..GCGGAATATG TTCAGTTCTC TATAGATTTG TTCAGTGTGG GTAAATCGGG  51  GGGCGGTATA CCTAAGGCTA AGCCTGTGTT TGATGCGAAA CCGAGATGGG 101  AGGTTGATAG GAAGCTTAAT AAATTGACAA CTCGTGAGCA GGTGGAGAAA 151  AATGTTCAGG AAACGAGAAG AAGGAGTCAG AGTAGTCAGT TTAAAGCCCA 201  TGCGCAACGA GAATGGGAAA ATAAAACAGG GTTAGATTTT AATCATTTTA 251  TAGGTGGTGA TATCAATAAA AAAGGCACAG TAACAGGAGG GCATAGTCTA 301  ACCCGTGGTG ATGTACGGGT GATACAACAA ACCTCGGCAC CTGATAAACA 351  TGGGGT.TTA TCAAGCGACA GTGGAAATTN A

This corresponds to the amino acid sequence <SEQ ID 458; ORF46>:

  1 ..AEYVQFSIDL FSVGKSGGGI PKAKPVFDAK PRWEVDRKLN KLTTREQVEK  51  NVQETRRRSQ SSQFKAHAQR EWENKTGLDF NHFIGGDINK KGTVTGGHSL 101  TRGDVRVIQQ TSAPDKHGXL SSDSGNX

Further work revealed further partial nucleotide sequence <SEQ ID 459>:

  1 ..GCAGTGTGCC TnCCGATGCA TGCACACGCC TCAnATTTGG CAAACGATTC  51  TTTTATCCGG CAGGTTCTCG ACCGTCAGCA TTTCGAACCC GACGGGAAAT 101  ACCACCTATT CGGCAGCAGG GGGGAACTTG CCGAGCGCCA GTCTCATATC 151  GGATTGGGAA AAATACAAAG CCATCAGTTG GGCAACCTGA TGATTCAACA 201  GGCGGCCATT AAAGGAAATA TCGGCTACAT TGTCCGCTTT TCCGATCACG 251  GGCACGAAGT CCATTCCCCs TTCGACAACC ATGCCTCACA TTCCGATTCT 301  GATGAAGCCG GTAGTCCCGT TGACGGATTT AGCCTTTACC GCATCCATTG 351  GGACGGATAC GAACACCATC CCGCCGACGG CTATGACGGG CCACAGGGCG 401  GCGGCTATCC CGCTCCCAAA GGCGCGAGGG ATATATACAG TTACGACATA 451  AAAGGCGTTG CCCAAAATAT CCGCCTCAAC CTGACCGACA ACCGCAGCAC 501  CGGACAACGG CTTGCCGACC GTTTCCACAA TGCCGGTAGT ATGCTGACGC 551  AAGGAGTAGG CGACGGATTC AAACGCGCCA CCCGATACAG CCCCGAGCTG 601  GACAGATCGG GCAATGCCGC CGAAGCCTTC AACGGCACTG CAGATATCGT 651  TAAAAACATC ATCGGCGCTG CAGGAGAAAT TGT

This corresponds to the amino acid sequence <SEQ ID 460; ORF46-1>:

  1 ..AVCLPMHAHA SXLANDSFIR QVLDRQHFEP DGKYHLFGSR GELAERQSHI  51  GLGKIQSHQL GNLMIQQAAI KGNIGYIVRF SDHGHEVHSP FDNHASHSDS 101  DEAGSPVDGF SLYRIHWDGY EHHPADGYDG PQGGGYPAPK GARDIYSYDI 151  KGVAQNIRLN LTDNRSTGQR LADRFHNAGS MLTQGVGDGF KRATRYSPEL 201  DRSGNAAEAF NGTADIVKNI IGAAGEI

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. gonorrhoeae

ORF46 shows 98.2% identity over, a 111 aa overlap with a predicted ORF(ORF46ng) from N. gonorrhoeae:

A partial ORF46ng nucleotide sequence <SEQ ID 461> is predicted toencode a protein having partial amino acid sequence <SEQ ID 462>:

  1 ..RRLKHCCHAR LGSAFHRKQD GAHQRFGRYG ATQRLCRSSH PRLGSPKPQC  51  RTRHRSRQQY LYGSHPHQRD WSCPGKIQLG RHHGTSCRAV ADXRDRICER 101  EIRRQRQXCR CRLGKIPSLS IPKYPLKLEQ RYGKENITSS TVPPSNGKNV 151  KLADQRHPKT GVPFDGKGFP NFEKHVKYDT KLDIQELSGG GIPKAKPVFD 201  AKPRWEVDRK LNKLTTREQV EKNVQETRRR SQSSQFKAHA QREWENKTGL 251  DFNHFIGGDI NKKGAVTGGH SLTRGDVRVI QQTSAPDKHG VLSSDSGN*

Further work revealed the complete gonococcal DNA sequence <SEQ ID 463>:

1 TTGGGCATTT CCCGCAAAAT ATCCCTTATT CTGTCCATAC TGGCAGTGTG 51 CCTGCCGATGCATGCACACG CCTCAGATTT GGcaAACGAT CCCTTTATCC 101 GgCaggttcT CGaccGTCAGCATTTCGaac ccgacggGAa ATACCaCCTA 151 TTcggCaGCA GGGGGGAGCT TgccnagcGCaacggccATa tcggattggG 201 aaacaTAcaa Agccatcagt tGggccacct gatgattcaacaggcggccg 251 ttgaaggaaA TAtcgGctac attgtccgct tttccgatca cgggcacaaa301 ttccattcgc ccttcGAcaa ccaTGCCTCA CATTCCGATT CTGACGAAGC 351CGGTAGTCCC GTTGACGGAT TCAGCCTTTA CCGCATCCAT TGGGACGGAT 401 ACGAACACCATCCCGCCGAC GGCTATGACG GGCCACAGGG CGGCGGCTAT 451 CCCGCTCCCA AAGGCGCGAGGGATATATAC AGCTACGACA TAAAAGGCGT 501 TGCCCAAAAT ATCCGCCTCA ACCTGACCGACAACCGCAGC ACCGGACAAC 551 GGCTTGCCGA CCGTTTCCAC AATGCCGGCG CTATGCTGACGCAAGGAGTA 601 GGCGACGGAT TCAAACGCGC CACCCGATAC AGCCCCGAGC TGGACAGATC651 GGGCAATGCc gccGAAGCCT TCAACGGCAC TGCAGATATC GTCAAAAACA 701TCATCGGCGC GGCAGGAGAA ATTGTCGGCG CAGGCGATGC CGTGCagGGT 751 ATAAGCGAAGGCTCAAACAT TGCTGTCATG CACGGCTTGG GTCTGCTTTC 801 CACCGAAAAC AAGATGGCGCGCATCAACGA TTTGGCAGAT ATGGCGCAAC 851 TCAAAGACTA TGCCGCAGCA GCCATCCGCGATTGGGCAGT CCAAAACCCC 901 AATGCCGCAC AAGGCATAGA AGCCGTCAGC AATATCTTTATGGCAGCCAT 951 CCCCATCAAA GGGATTGGAG CTGTCCGGGG AAAATACGGC TTGGGCGGCA1001 TCACGGCACA TCCTGTCAAG CGGTCGCAGA TGGGCGCGAT CGCATTGCCG 1051AAAGGGAAAT CCGCCGTCAG CGACAATTTT GCCGATGCGG CATACGCCAA 1101 ATACCCGTCCCCTTACCATT CCCGAAATAT CCGTTCAAAC TTGGAGCAGC 1151 GTTACGGCAA AGAAAACATCACCTCCTCAA CCGTGCCGCC GTCAAACGGC 1201 AAAAATGTCA AACTGGCAGA CCAACGCCACCCGAAGACAG GCGTACCGTT 1251 TGACGGTAAA GGGTTTCCGA ATTTTGAGAA GCACGTGAAATATGATACGA 1301 AGCTCGATAT TCAAGAATTA TCGGGGGGCG GTATACCTAA GGCTAAGCCT1351 GTGTTTGATG CGAAACCGAG ATGGGAGGTT GATAGGAAGC TTAATAAATT 1401GACAACTCGT GAGCAGGTGG AGAAAAATGT TCAGGAAACG AGAAGAAGGA 1451 GTCAGAGTAGTCAGTTTAAA GCCCATGCGC AACGAGAATG GGAAAATAAA 1501 ACAGGGTTAG ATTTTAATCATTTTATAGGT GGTGATATCA ATAAGAAAGG 1551 CACAGTAACA GGAGGGCATA GTCTAACCCGTGGTGATGTA CGGGTGATAC 1601 AACAAACCTC GGCACCTGAT AAACATGGGG TTTATCAAGCGACAGTGGAA 1651 ATTAAAAAGC CTGATGGAAG TTGGGAGGTG AAAACGAAAA AAGGTGGGAA1701 AGTGATGACC AAGCACACCA TGTTCCCAAA AGATTGGGAT GAGGCTAGAA 1751TTAGGGCTGA AGTTACTTCG GCTTGGGAAA GTAGAATAAT GCTTAAGGAT 1801 AATAAATGGCAGGGTACAAG TAAATCGGGT ATTAAAATAG AAGGATTTAC 1851 CGAACCTAAT AGAACAGCATATCCCATTTA TGAATAG

This corresponds to the amino acid sequence <SEQ ID 464; ORF46ng-1>:

  1 LGISRKISLI LSILAVCLPM HAHASDLAND PFIRQVLDRQ HFEPDGKYHL  51FGSRGELAXR NGHIGLGNIQ SHQLGHLMIQ QAAVEGNIGY IVRFSDHGHK 101 FHSPFDNHASHSDSDEAGSP VDGFSLYRIH WDGYEHHPAD GYDGPQGGGY 151 PAPKGARDIY SYDIKGVAQNIRLNLTDNRS TGQRLADRFH NAGAMLTQGV 201 GDGFKRATRY SPELDRSGNA AEAFNGTADIVKNIIGAAGE IVGAGDAVQG 251 ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAQLKDYAAAAIRDWAVQNP 301 NAAQGIEAVS NIFMAAIPIK GIGAVRGKYG LGGITAHPVK RSQMGAIALP351 KGKSAVSDNF ADAAYAKYPS PYHSRNIRSN LEQRYGKENI TSSTVPPSNG 401KNVKLADQRH PKTGVPFDGK GFPNFEKHVK YDTKLDIQEL SGGGIPKAKP 451 VFDAKPRWEVDRKLNKLTTR EQVEKNVQET RRRSQSSQFK AHAQREWENK 501 TGLDFNHFIG GDINKKGTVTGGHSLTRGDV RVIQQTSAPD KHGVYQATVE 551 IKKPDGSWEV KTKKGGKVMT KHTMFPKDWDEARIRAEVTS AWESRIMLKD 601 NKWQGTSKSG IKIEGFTEPN RTAYPIYE*

ORF46ng-1 and ORF46-1 show 94.7% identity in 227 aa overlap:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF46ng-1 shows 87.4% identity over a 486aa overlap with an ORF (ORF46a)from strain A of N. meningitidis:

The complete length ORF46a DNA sequence <SEQ ID 465> is:

1 TTGGGCATTT CCCGCAAAAT ATCCCTTATT CTGTCCATAC TGGCAGTGTG 51 CCTGCCGATGCATGCACACG CCTCAGATTT GGCAAACGAT TCTTTTATCC 101 GGCAGGTTCT CGACCGTCAGCATTTCGAAC CCGACGGGAA ATACCACCTA 151 TTCGGCAGCA GGGGGGAACT TGCCGAGCGCAGCGGTCATA TCGGATTGGG 201 AAACATACAA AGCCATCAGT TGGGCAACCT GTTCATCCAGCAGGCGGCCA 251 TTAAAGGAAA TATCGGCTAC ATTGTCCGCT TTTCCGATCA CGGGCACGAA301 GTCCATTCCC CCTTCGACAA CCATGCCTCA CATTCCGATT CTGATGAAGC 351CGGTAGTCCC GTTGACGGAT TCAGCCTTTA CCGCATCCAT TGGGACGGAT 401 ACGAACACCATCCCGCCGAC GGCTATGACG GGCCACAGGG CGGCGGCTAT 451 CCCGCTCCCA AAGGCGCGAGGGATATATAC AGCTACGACA TAAAAGGCGT 501 TGCCCAAAAT ATCCGCCTCA ACCTGACCGACAACCGCAGC ACCGGACAAC 551 GGCTTGTCGA CCGTTTCCAC AATACCGGTA GTATGCTGACGCAAGGAGTA 601 GGCGACGGAT TCAAACGCGC CACCCGATAC AGCCCCGAGC TGGACAGATC651 GGGCAATGCC GCCGAAGCTT TCAACGGCAC TGCAGATATC GTCAAAAACA 701TCATCGGCGC GGCAGGAGAA ATTGTCGGCG CAGGCGATGC CGTGCAGGGT 751 ATAAGCGAAGGCTCAAACAT TGCTGTTATG CACGGCTTGG GTCTGCTTTC 801 CACCGAAAAC AAGATGGCGCGCATCAACGA TTTGGCAGAT ATGGCGCAAC 851 TCAAAGACTA TGCCGCAGCA GCCATCCGCGATTGGGCAGT CCAAAACCCC 901 AATGCCGCAC AAGGCATAGA AGCCGTCAGC AATATCTTTACGGCAGTCAT 951 CCCCGTCAAA GGGATTGGAG CTGTTCGGGG AAAATACGGC TTGGGCGGCA1001 TCACGGCACA TCCTGTCAAG CGGTCGCAGA TGGGCGAGAT CGCATTGCCG 1051AAAGGGAAAT CCGCCGTCAG CGACAATTTT GCCGATGCGG CATACGCCAA 1101 ATACCCGTCCCCTTACCATT CCCGAAATAT CCGTTCAAAC TTGGAGCAGC 1151 GTTACGGCAA AGAAAACATCACCTCCTCAA CCGTGCCGCC GTCAAACGGA 1201 AAGAATGTGA AACTGGCAAA CAAACGCCACCCGAAGACCA AAGTGCCGTT 1251 TGACGGTAAA GGGTTTCCGA ATTTTGAAAA AGACGTAAAATACGATACGA 1301 GAATTAATAC CGCTGTACCA CAAGTGAATC CTATAGATGA ACCCGTCTTT1351 AATCCTAAAG GTTCTGTCGG ATCGGCTCAT TCTTGGTCTA TAACTGCCAG 1401AATTCAATAC GCAAAATTAC CAAGGCAAGG TAGAATCAGA TATATCCCAC 1451 CTAAAAATTACTCTCCTTCA GCACCGCTAC CAAAAGGACC TAATAATGGA 1501 TATTTGGATA AATTTGGTAATGAATGGACT AAAGGTCCAT CAAGAACTAA 1551 AGGTCAAGAA TTTGAATGGG ATGTTCAATTGTCTAAAACA GGAAGAGAGC 1601 AACTTGGATG GGCTAGTAGG GATGGTAAGC ATTTAAATATATCAATTGAT 1651 GGAAAGATTA CACACAAATG A

This corresponds to the amino acid sequence <SEQ ID 466>:

  1 LGISRKISLI LSILAVCLPM HAHASDLAND SFIRQVLDRQ HFEPDGKYHL  51FGSRGELAER SGHIGLGNIQ SHQLGNLFIQ QAAIKGNIGY IVRFSDHGHE 101 VHSPFDNHASHSDSDEAGSP VDGFSLYRIH WDGYEHHPAD GYDGPQGGGY 151 PAPKGARDIY SYDIKGVAQNIRLNLTDNRS TGQRLVDRFH NTGSMLTQGV 201 GDGFKRATRY SPELDRSGNA AEAFNGTADIVKNIIGAAGE IVGAGDAVQG 251 ISEGSNIAVM HGLGLLSTEN KMARINDLAD MAQLKDYAAAAIRDWAVQNP 301 NAAQGIEAVS NIFTAVIPVK GIGAVRGKYG LGGITAHPVK RSQMGEIALP351 KGKSAVSDNF ADAAYAKYPS PYHSRNIRSN LEQRYGKENI TSSTVPPSNG 401KNVKLANKRH PKTKVPFDGK GFPNFEKDVK YDTRINTAVP QVNPIDEPVF 451 NPKGSVGSAHSWSITARIQY AKLPRQGRIR YIPPKNYSPS APLPKGPNNG 501 YLDKFGNEWT KGPSRTKGQEFEWDVQLSKT GREQLGWASR DGKHLNISID 551 GKITHK*

Based on this analysis, including the presence of a RGD sequence in thegonococcal protein, typical of adhesins, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 56

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 467>:

  1 ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC CGCCATTCCT  51GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTTGCC CCCAATGCGG 101 TGTTTTGGGTTTTGGCACTG CTGACCGCCA CCGCCCGCCC GATTGTCAAT 151 TTGGACTATC TTCCCGCCGCGCTGCTGATC GCCCTGCCTT GGCGTTTCGT 201 CAAAATTGCC GGCGTATTGG CGTTTTGGCTGGCGGTTTTG TTTGACGGGC 251 TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCATCGGCGCCATC 301 AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC AGATAATGAC351 CGGGCTG...

This corresponds to the amino acid sequence <SEQ ID 468; ORF48>:

  1 MNIHTLLSKQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL LTATARPIVN  51LDYLPAALLI ALPWRFVKIA GVLAFWLAVL FDGLMMVIQL FPFMDLIGAI 101 NLVPFILTAPAPYQIMTGL...

Further work revealed the complete nucleotide sequence <SEQ ID 469>:

1 ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC CGCCATTCCT 51 GCCGAAACGGCTGCTGCTGT CCCTGCTGAT ACTGCTTGCC CCCAATGCGG 101 TGTTTTGGGT TTTGGCACTGCTGACCGCCA CCGCCCGCCC GATTGTCAAT 151 TTGGACTATC TTCCCGCCGC GCTGCTGATCGCCCTGCCTT GGCGTTTCGT 201 CAAAATTGCC GGCGTATTGG CGTTTTGGCT GGCGGTTTTGTTTGACGGGC 251 TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCAT CGGCGCCATC301 AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC AGATAATGAC 351CGGGCTGTTG CTGCTGTATA TGCTGGCGAT GCCGTTTGTG TTGCAGAAAG 401 CCGCCGCCAAAACCGACTTC CGGCACATTG CCGTCTGCGC CGCCGTTGTG 451 GCGGCAGCCG GCTATTTCACCGGCCATTTG AGTTACTACG ACCGGGGTCG 501 GATGGCCAAT ATCTTCGGCG CAAACAACTTCTACTACGCC AAAAGTCAGG 551 CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTACCGCCGGCCTG 601 GTCGATCCCG TCTTCCTCCC CTTGGGCAAT CAACAGCGTG CCGCCACGCA651 TCTGAACGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC GCCGAATCTT 701GGGGGCTGCC GGCCAATCCC GAACTTCAAA ACGCCACTTT TGCCAAACTG 751 CTGGCGCAAAAAGACCGTTT TTCGGTTTGG GAAAGCGGCA GTTTTCCCTT 801 CATCGGCGCG ACGGTCGAAGGCGAAATGCG CGAACTGTGT GCCTACGGCG 851 GTTTGCGCGG GTTCGCACTG CGCCGCGCGCCCGACGAAAA ATTTGCCCGC 901 TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCTTTGCGATGCA 951 CGGCGCGGGC AGTTCGCTTT ACGACCGCTT CAGCTGGTAT CCGAGGGCGG1001 GCTTTCAAGA AATCAAAACC GCCGAAAACC TGATCGGTAA AAAAACCTGC 1051GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG AAGTGTCGGC 1101 ATTTTTCAAAAAACACGACA AGGGACTGTT TTACTGGATG ACGCTGACCA 1151 GCCACGCCGA CTATCCCGAATCCGACATTT TCAACCACAG GCTCAAATGC 1201 ACCGAATATG GCCTGCCCGC CGAAACCGACCTCTGCCGCA ATTTCAGCCT 1251 GCACACCCAA TTCTTCGACC AACTGGCGGA TTTGATCCAACGCCCCGAAA 1301 TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC GCCCGTCGGC1351 AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGGCACG TCGCCTGGCT 1401GAACTTCAAA ATCAAATAA

This corresponds to the amino acid sequence <SEQ ID 470; ORF48-1>:

  1 MNIHTLLSKQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL LTATARPIVN  51LDYLPAALLI ALPWRFVKIA GVLAFWLAVL FDGLMMVIQL FPFMDLIGAI 101 NLVPFILTAPAPYQIMTGLL LLYMLAMPFV LQKAAAKTDF RHIAVCAAVV 151 AAAGYFTGHL SYYDRGRMANIFGANNFYYA KSQAMLYTVS QNADFITAGL 201 VDPVFLPLGN QQRAATHLNE PKSQKILFIVAESWGLPANP ELQNATFAKL 251 LAQKDRFSVW ESGSFPFIGA TVEGEMRELC AYGGLRGFALRRAPDEKFAR 301 CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQEIKT AENLIGKKTC351 AIFGGVCDSE LFGEVSAFFK KHDKGLFYWM TLTSHADYPE SDIFNHRLKC 401TEYGLPAETD LCRNFSLHTQ FFDQLADLIQ RPEMKGTEVI IVGDHPPPVG 451 NLNETFRYLKQGHVAWLNFK IK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF48 shows 94.1% identity over a 119aa overlap with an ORF (ORF48a)from strain A of N. meningitidis:

The complete length ORF48a nucleotide sequence <SEQ ID 471> is:

1 ATGAATATTC ACACCCTGCT CTCCAAACAA TGGACGCTGC CGCCATTCCT 51 GCCGAAACGGCTGCTGCTGT CCCTGCTGAT ACTGCTNNCC CCCAATGCGG 101 TGTTTTGGGT TTTGGCACTGCTGACCGCCA CCGCCCGCCC GATTGTCAAT 151 TTGGANTACC TTCCCGCCGC GCTGCTGATCGCCCTGCCTT GGCGTNTCGT 201 CAAAATTGNC GGCGTATTGG CGTNTTGGCT GGCGGTTTTGTTTGACGGGC 251 TGATGATGGT GATCCAACTC TTCCCTTTTA TGGATCTCAT CGGCGCCATC301 AACCTCGTCC CCTTCATCNT GACCGCCCCC GCCCTTTATC AGATAATGAC 351CGGGCTGTTA CTGCTGTATA TGCTGGCGAT GCCGTTTGTG TTGCAGAAAG 401 CCGCCGCCAAAACCGACTTC CGACACATTG CCGCCTGTGC CGCCGTTGTG 451 GTGGCAGCCG GCTATTTTACCGGCCATTTG AGTTANTACG ACCGGGGGCG 501 GATGGCCAAT ATCTTCGGCG CAAACAACTTCTATTACGCC AAAAGTCAGG 551 CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTACCGCCGGCCTG 601 GTCGATCCCG TCTTCCTCCC CTTGGGCAAT CAACAGCGTG CCGCCACGCA651 TCTGAACGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC GCCGAATCTT 701GGGGGCTGCC GGCCAATCCC GAACTTCAAA ACGCCACTTT TGCCAAACTG 751 CTGGCGCAAAAAGANCGTTT TTCGGTTTGG GAAAGCGGCA GTTTTCCCTT 801 CATCGGCGCG ACGATCGAAGGCGAAATGCG CGAACTGTGT GCCTACGGCG 851 GTTTGCGCGG GTTCGCACTG CGCCGCGCGCCCGACGAAAA ATTTGCCCGC 901 TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCTTTGCGATGCA 951 CGGCGCGGGC AGTTCGCTTT ACGACCGCTT CAGCTGGTAT CCGAGGGCGG1001 GCTTTCAAGA AATCAAAACC GCCGAAAACC TGATCGGTAA AAAAACCTGC 1051GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG AAGTGTCGGC 1101 ANTTTTCAAAAAACACGACA AGGGACTGTT TTACTGGATG ACGCTGACCA 1151 GCCACGCCGA CTATCCCGAATCNGACATTT TCAACCACAG GCTCAAATGC 1201 ACCGAATATG GCCTGCCCGC CGAAACCGACNTCTGCCGCA ATTTCAGCCT 1251 GCACACCCAA TTCTTCGACC AACTGGCGGA TTTGATCCAACGCCCCGAAA 1301 TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC GCCCGTCGGC1351 AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGGCACG TCGNCTGGCT 1401GAACTTCAAA ATCAAATAA

This encodes a protein having amino acid sequence <SEQ ID 472>:

  1 MNIHTLLSKQ WTLPPFLPKR LLLSLLILLX PNAVFWVLAL LTATARPIVN  51LXYLPAALLI ALPWRXVKIX GVLAXWLAVL FDGLMMVIQL FPFMDLIGAI 101 NLVPFIXTAPALYQIMTGLL LLYMLAMPFV LQKAAAKTDF RHIAACAAVV 151 VAAGYFTGHL SXYDRGRMANIFGANNFYYA KSQAMLYTVS QNADFITAGL 201 VDPVFLPLGN QQRAATHLNE PKSQKILFIVAESWGLPANP ELQNATFAKL 251 LAQKXRFSVW ESGSFPFIGA TIEGEMRELC AYGGLRGFALRRAPDEKFAR 301 CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQEIKT AENLIGKKTC351 AIFGGVCDSE LFGEVSAXFK KHDKGLFYWM TLTSHADYPE SDIFNHRLKC 401TEYGLPAETD XCRNFSLHTQ FFDQLADLIQ RPEMKGTEVI IVGDHPPPVG 451 NLNETFRYLKQGHVXWLNFK IK*

ORF48a and ORF48-1 show 96.8% identity in 472 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF48 shows 97.5% identity over a 119aa overlap with a predicted ORF(ORF48ng) from N. gonorrhoeae:

The ORF48ng nucleotide sequence <SEQ ID 473> was predicted to encode aprotein having amino acid sequence <SEQ ID 474>:

  1 MNIHALLSEQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL LTATARPIVN  51LDYLPAALLI ALPWRFVKIA GVLAFWPAVL FDGLMMVIQL FPFMDLIGAI 101NLVPFILTAP APYQIMTGLL LLYMLAMPFV LQKAAVKTDF RHIAVCAAVV 151AAARYFTGPF ELLRTGGRWQ YVQHRRLLLS GSRASFRRRQ KADVLRRLGN 201 PYASMGNGG..

Further work identified the complete gonococcal DNA sequence <SEQ ID475>:

1 ATGAATATTC ACGCCCTGCT CTCCGAACAA TGGACGCTGC CGCCATTCCT 51GCCGAAACGG CTGCTGCTGT CCCTGCTGAT ACTGCTGGCC CCCAATGCGG 101TGTTTTGGGT TTTGGCACTG CTGACCGCCA CCGCCCGCCC GATTGTCAAT 151TTGGACTACC TTCCCGCCGC GCTGCTGATC GCCCTGCCTT GGCGTTTCGT 201CAAAATTGCC GGCGTATTGG CGTTTTGGCC GGCGGTTTTG TTTGACGGGC 251TGATGATGGT GATCCAACTC TTCCCTTTTA TGGACCTCAT CGGCGCCATC 301AACCTCGTCC CCTTCATCCT GACCGCCCCC GCCCCTTATC AGATAATGAC 351CGGGCTGTTG CTGCTGTATA TGCTGGCGAT GCCGTTTGTG TTGCAAAAAG 401CCGCCGTCAA AACCGACTTC CGACACATTG CCGTCTGTGC CGCCGTTGTG 451GCGGCAGCCG GCTATTTCAC CGGCCATTTG AGTTACTACG ACCGGGGGCG 501GATGGCCAAT ATCTTCGGCG CAAACAACTT CTATTACGCc aAAAGTCAGG 551CGATGCTCTA CACCGTCAGC CAGAATGCCG ACTTTATTAC CGCCGgcctG 601GTCGACCCCG TCTTCCTCCC CTTGGGCAAT CAGCAGCGTG CCGCCACGCG 651GCTGAGTGAG CCGAAATCTC AAAAAATCCT CTTTATCGTC GCCGAATCTT 701GGGGGCTGCC GGGCAATCCC GAGCTTCAAA ACGCCACTTT TGCCAAACTG 751CTGGCGCAAA AAGACCGTTT TTCGGTTTGG GAAAGCGGCA GTTTTCCCTT 801CATCGGCGCG ACGGTCGAAG GCGAAATGCG CGAATTGTGC GCCTACGGCG 851GTTTGCGCGG GTTCGCACTG CGCCGCGCGC CCGACGAAAA ATTTGCCCGC 901TGCCTCCCCA ACCGTTTGAA ACAAGAAGGT TACGCCACCT TTGCGATGCA 951CGGCGCGGGT AGTTCGCTTT ACGACCGCTT CAGCTGGTAT CCGAGGGCGG 1001GCTTTCAAAA AATCAAAACC GCCGAAAACC TGATCGGTAA AAAAACCTGC 1051GCCATTTTCG GCGGCGTGTG CGACAGCGAG CTGTTCGGCG AAGTGTCGGC 1101ATTTTTCAAA AAACACGACA AGGGACTGTT TTACTGGATG ACGCTGACCA 1151GCCACGCCGA CTATCCCGAA TCCGACATTT TCAACCACAG GCTCAAATGC 1201ACCGAATACG GCCTGCCCGC CGAAACCGAC CTCTGCCGCA ATTTCAGCCT 1251GCACACCCAA TtcttcgACC AACTGGCGGA TTTGATCCGA CGCCCCGAAA 1301TGAAAGGCAC GGAAGTCATC ATCGTCGGCG ACCATCCGCC GCCCGTCGGC 1351AACCTCAATG AAACCTTCCG CTACCTCAAA CAGGGACACG TCGCCTGGCT 1401GCACTTCAAA ATCAAATAA

This encodes a protein having amino acid sequence <SEQ ID 476;ORF48ng-1>:

  1 MNIHALLSEQ WTLPPFLPKR LLLSLLILLA PNAVFWVLAL LTATARPIVN  51LDYLPAALLI ALPWRFVKIA GVLAFWPAVL FDGLMMVIQL FPFMDLIGAI 101NLVPFILTAP APYQIMTGLL LLYMLAMPFV LQKAAVKTDF RHIAVCAAVV 151AAAGYFTGHL SYYDRGRMAN IFGANNFYYA KSQAMLYTVS QNADFITAGL 201VDPVFLPLGN QQRAATRLSE PKSQKILFIV AESWGLPGNP ELQNATFAKL 251LAQKDRFSVW ESGSFPFIGA TVEGEMRELC AYGGLRGFAL RRAPDEKFAR 301CLPNRLKQEG YATFAMHGAG SSLYDRFSWY PRAGFQKIKT AENLIGKKTC 351AIFGGVCDSE LFGEVSAFFK KHDKGLFYWM TLTSHADYPE SDIFNHRLKC 401TEYGLPAETD LCRNFSLHTQ FFDQLADLIR RPEMKGTEVI IVGDHPPPVG 451NLNETFRYLK QGHVAWLHFK IK*

ORG48ng-1 and ORF48-1 show 97.9% identity in 472 aa overlap:

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and two putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 57

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 477>:

  1 ..GTGAGCGGAC GTTACCGCGC TTTGGATCGC GTTTCCAAAA TCATCATCGT  51  TACTTTGAGT ATCGCCACGC TTGCCGCCGC CGGCATCGCT ATGTCGCGCG 101  GTATGCAGAT GCAGTCCGAT TTTATCGAGC CGACACCGTG GACGCTTGCC 151  GGTTTGGGCT TCCTGATCGC GCTGATGGGC TGGATGCCCG CGCCGATTGA 201  AATTTCCGCC ATCAATTCTT TGTGGGTAAC CGAAAAACAA CGCATCAATC 251  CTTCCGAATA CCGCGACGGG ATTTTTGAAT TCAACGTCGG TTATATCGCC 301  AGTGCGGTTT TGGCTTTGGT TTTCCTTGCA CTGGGCGC.G TAGCGCCGAA 351  CGGCAACGGC GA.ACAGTGC AGATGGCGGG CGGCAAATAT AACGGGCAAT 401  TGATCAATAT GTACGCC..

This corresponds to the amino acid sequence <SEQ ID 478; ORF53>:

  1 ..VSGRYRALDR VSKIIIVTLS IATLAAAGIA MSRGMQMQSD FIEPTPWTLA  51  GLGFLIALMG WMPAPIEISA INSLWVTEKQ RINPSEYRDG IFEFNVGYIA 101  SAVLALVFLA LGXVAPNGNG XTVQMAGGKY NGQLINMYA..

Further work revealed the complete nucleotide sequence <SEQ ID 479>:

1 ATGTCCGAAC AACATATTTC GACTTGGAAA AGTAAAATCA ACGCATTGGG 51TCCGGGGATC ATGATGGCTT CGGCGGCGGT CGGCGGTTCG CACCTGATTG 101CCTCGACGCA GGCGGGCGCG CTTTACGGCT GGCAGATCGC GCTCATCATC 151ATCCTGACCA ACCTCTTCAA ATACCCGTTT TTCCGCTTCA GCGCGCATTA 201CACGCTGGAC ACGGGCAAGA GCCTGATTGA AGGTTATGCC GAGAAAAGCC 251GCGTTTATTT GTGGGTATTC CTGATTTTGT GCATCCTCTC CGCCACGATT 301AACGCGGGCG CGGTCGCCAT TGTAACCGCC GCCATCGTCA AAATGGCGAT 351TCCCTCGCTG ATGTTTGATG CCGGCACGGT TGCCGCCTTG ATTATGGCAT 401CCTGCCTGAT TATTTTGGTG AGCGGACGTT ACCGCGCTTT GGATCGCGTT 451TCCAAAATCA TCATCGTTAC TTTGAGTATC GCCACGCTTG CCGCCGCCGG 501CATCGCTATG TCGCGCGGTA TGCAGATGCA GTCCGATTTT ATCGAGCCGA 551CACCGTGGAC GCTTGCCGGT TTGGGCTTCC TGATCGCGCT GATGGGCTGG 601ATGCCCGCGC CGATTGAAAT TTCCGCCATC AATTCTTTGT GGGTAACCGA 651AAAACAACGC ATCAATCCTT CCGAATACCG CGACGGGATT TTTGATTTCA 701ACGTCGGTTA TATCGCCAGT GCGGTTTTGG CTTTGGTTTT CCTTGCACTG 751GGCGCGTTTG TGCAATACGG CAACGGCGAA GCAGTGCAGA TGGCGGGCGG 801CAAATATATC GGGCAATTGA TCAATATGTA CGCCGTTACC ATCGGCGGCT 851GGTCGCGCCC GCTGGTGGCG TTTATCGCGT TTGCCTGTAT GTACGGCACG 901ACGATTACCG TCGTGGACGG CTATGCCCGT GCCATTGCCG AACCCGTGCG 951CCTGCTGCGC GGAAAAGACA AAACGGGCAA CGCCGAATTC TTTGCCTGGA 1001ATATTTGGGT GGCGGGCAGC GGTTTGGCGG TGATTTTCTG GTTTGACGGC 1051GTAATGGCGA ATCTGCTCAA ATTTGCGATG ATTGCCGCTT TTGTGTCCGC 1101CCCTGTGTTT GCCTGGCTGA ATTACCGTTT GGTTAAAGGT GATGAAAAAC 1151ACAAACTCAC ATCAGGTATG AATGCCCTTG CATTGGCAGG CTTGATTTAT 1201CTGACCGGTT TTACCGTTTT GTTCTTATTG AATTTGGCGG GAATGTTCAA 1251 ATGA

This corresponds to the amino acid sequence <SEQ ID 480; ORF53-1>:

  1 MSEQHISTWK SKINALGPGI MMASAAVGGS HLIASTQAGA LYGWQIALII  51ILTNLFKYPF FRFSAHYTLD TGKSLIEGYA EKSRVYLWVF LILCILSATI 101NAGAVAIVTA AIVKMAIPSL MFDAGTVAAL IMASCLIILV SGRYRALDRV 151SKIIIVTLSI ATLAAAGIAM SRGMQMQSDF IEPTPWTLAG LGFLIALMGW 201MPAPIEISAI NSLWVTEKQR INPSEYRDGI FDFNVGYIAS AVLALVFLAL 251GAFVQYGNGE AVQMAGGKYI GQLINMYAVT IGGWSRPLVA FIAFACMYGT 301TITVVDGYAR AIAEPVRLLR GKDKTGNAEF FAWNIWVAGS GLAVIFWFDG 351VMANLLKFAM IAAFVSAPVF AWLNYRLVKG DEKHKLTSGM NALALAGLIY 401LTGFTVLFLL NLAGMFK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF53 shows 93.5% identity over a 139aa overlap with an ORF (ORF53a)from strain A of N. meningitidis:

The complete length ORF53a nucleotide sequence <SEQ ID 481> is:

1 ATGTCCGAAC AACATATTTC GACTTGGAAA AGTAAAATCA ACGCATTGGG 51ACCGGGGATT ATGATGGCTT CGGCGGCGGT CGGCGGTTCG CACCTGATTG 101CCTCGACGCA GGCGGGCGCG CTTTACGGCT GGCAGATCGC GCTCATCATC 151ATCCTGACCA ACCTCTTCAA ATACCCGTTT TTCCGCTTCA GCGCGCATTA 201CACGCTGGAC ACGGGCAAGA GCCTGATTGA AGGTTATGCC GAGAAAAGCC 251GCGTTTATTT GTGGGTATTC CTGATTTTGT GCATCCTCTC CGCCACGATT 301AACGCGGGCG CGGTCGCCAT TGTAACCGCC GCCATCGTCA AAATGGCGAT 351TCCCTCGCTG ATGTTTGATG CCGGCACGGT TGCCGCCTTG ATTATGGCAT 401CCTGCCTGAT TATTTTGGTG AGCGGACGTT ACCGCGCTTT GGATCGCGTT 451TCCAAAATCA TCATCGTTAC TTTGAGTATC GCCACGCTTG CCGCCGCCGG 501CATCGCTATG TCGCGCGGTA TGCAGATGCA GTCCGATTTT ATCGAGCCGA 551CACCGTGGAC GCTTGCCGGT TTGGGCTTCC TGATCGCGCT GATGGGCTGG 601ATGCCCGCGC CGATTGAAAT TTCCGCCATC AATTCTTTGT GGGTAACCGA 651AAAACAACGC ATCAATCCTT CCGAATACCG CGACGGGATT TTTGATTTCA 701ACGTCGGTTA TATCGCCAGT GCGGTTTTGG CTTTGGTTTT CCTTGCACTG 751GGCGCGTTTG TGCAATACGG CAACGGCGAA GCAGTGCAGA TGGCGGGCGG 801CAAATATATC GGGCAATTGA TCAATATGTA CGCCGTTACC ATCGGCGGCT 851GGTCGCGCCC GCTGGTGGCG TTTATCGCGT TTGCCTGTAT GTACGGCACG 901ACGATTACCG TTGTGGACGG CTATGCCCGT GCCATTGCCG AACCCGTGCG 951CCTGCTGCGC GGAAAAGACA AAACGGGCAA CGCCGAATTC TTTGCCTGGA 1001ATATTTGGGT GGCGGGCAGC GGTTTGGCGG TGATTTTCTG GTTTGACGGC 1051GTAATGGCGA ATCTGCTCAA ATTTGCGATG ATTGCCGCTT TTGTGTCCGC 1101CCCTGTGTTT GCCTGGCTGA ATTACCGTTT GGTCAAAGGT GATGAAAAAC 1151ACAAACTCAC ATCAGGTATG AATGCCCTTG CATTGGCAGG CTTGATTTAT 1201CTGACCGGTT TTACCGTTTT GTTCTTATTG AATTTGGCGG GAATGTTCAA 1251 ATGA

This encodes a protein having amino acid sequence <SEQ ID 482>:

  1 MSEQHISTWK SKINALGPGI MMASAAVGGS HLIASTQAGA LYGWQIALII  51ILTNLFKYPF FRFSAHYTLD TGKSLIEGYA EKSRVYLWVF LILCILSATI 101NAGAVAIVTA AIVKMAIPSL MFDAGTVAAL IMASCLIILV SGRYRALDRV 151SKIIIVTLSI ATLAAAGIAM SRGMQMQSDF IEPTPWTLAG LGFLIALMGW 201MPAPIEISAI NSLWVTEKQR INPSEYRDGI FDFNVGYIAS AVLALVFLAL 251GAFVQYGNGE AVQMAGGKYI GQLINMYAVT IGGWSRPLVA FIAFACMYGT 301TITVVDGYAR AIAEPVRLLR GKDKTGNAEF FAWNIWVAGS GLAVIFWFDG 351VMANLLKFAM IAAFVSAPVF AWLNYRLVKG DEKHKLTSGM NALALAGLIY 401LTGFTVLFLL NLAGMFK*

ORF 53a shows 100.0% identity in 417 aa overlap with ORF53-1:

Homology with a Predicted ORF from N. gonorrhoeae

ORF53 shows 92.1% identity over a 139aa overlap with a predicted ORF(ORF53ng) from N. gonorrhoeae:

An ORF53ng nucleotide sequence <SEQ ID 483> was predicted to encode aprotein having amino acid sequence <SEQ ID 484>:

  1 MPKKSCVYLW VFLILCIASA TINAGAVAIV TAAIVKMAIP SLMFDAGTVA  51ALIMASCLII LVSGRYRALD RVSKIIIVTL SIATLAAAGI AMSRGMQMQP 101DFIEPTPWTL AGLGFLIALM GWMPAPIEIS AINSLWVTEK QRINPSEYRD 151GIFDFNVGYI ASAVLALVFL ALGAFVQYGN GEAVQMGGGK YIGQLINMYA 201VTIGGGSRPL VAFIAFACMY GAASTVVDGY ARAIAEPVRL LRGKDKTARP 251IVLLEKLGGR HRFGRDFLV*

Further analysis revealed further partial DNA gonococcal sequence <SEQID 485>:

1 ..aagaAAAGCT GCGTTTATTT GTGGGTTTTT TTGATTTTGT GTATCGCCTC 51  CGCCACGATT AACGCGGGCG CGGTCGCCAT TGTAACCGCC GCCATCGTCA 101  AAATGGCGAT TCCCTCGCTG ATGTTTGATG CCGGCACGGT TGCCGCCTTG 151  ATTATGGCAT CCTGCCTGAT TATTTTGGTG AGCGGACGTT ACCGCGCTTT 201  GGATCGTGTT TCCAAAATCA TCATTGTTAC TTTGAGCATC GCCACGCTTG 251  CCGCCGCCGG CATCGCTATG TCGCGCGGTA TGCAGATGCA GCCCGATTTT 301  ATCGAGCCGA CACCGTGGAC GCTTGCCGGT TTGGGCTTCC TGATCGCGCT 351  GATGGGCTGG ATGCCCGCGC CGATCGAAAT TTCCGCCATC AATTCTTTGT 401  GGGTAACCGA AAAACAACGC ATCAATCCTT CTGAATACCG CGACGGGATT 451  TTCGATTTCA ACGTCGGTTA TATCGCcagT GCGGTTTTGG CTTTGGTTTT 501  CCTTGCACTG GGCGCGTTTG TGCAATACGG CAACGGCGAA GCAGTGCAGA 551  TGGCGGGCGG CAAATATATC GGGCAATTGA TTAATATGTA TGCCGTAACC 601  ATCGGCGGCT GGTCTCGTCC GCTGGTGGCG TTTATCGCGT TTGCCTGTAT 651  GTACGGCACG ACGATTACCG TTGTGGACGG TTATGCGCGT GCCATTGCCG 701  AACCCGTGCG CCTGCTGCGC GGCAGGGATA AAACCGGCAA CGCCGAGTTG 751  TTtgccTGGA ATATTTGGGT GGCGGGCAGC GGTTTGGCGG TGATTTTCTG 801  GTTTGACggc gcaaTGGCgG AACtgcTCAA ATTTGCGATG ATtgccgcCT 851  TTGTGTCCGC CCCTGTGTTC GCCTGGCTCA ACTACCGCCT CGTCAAAGGG 901  GACAAACGCC ACAGGCTTAC CGCCGGTATG AACGCCCTTG CCATTGTCGG 951  CCTGCTCTAC CTGGCCGGGT TTGCCGTTTT GTTCCTGTTG AACCTTACCG 1001  GACTTTTGGC ATAG

This corresponds to the amino acid sequence <SEQ ID 486; ORF53ng-1>:

  1 ..KKSCVYLWVF LILCIASATI NAGAVAIVTA AIVKMAIPSL MFDAGTVAAL  51  IMASCLIILV SGRYRALDRV SKIIIVTLSI ATLAAAGIAM SRGMQMQPDF 101  IEPTPWTLAG LGFLIALMGW MPAPIEISAI NSLWVTEKQR INPSEYRDGI 151  FDFNVGYIAS AVLALVFLAL GAFVQYGNGE AVQMAGGKYI GQLINMYAVT 201  IGGWSRPLVA FIAFACMYGT TITVVDGYAR AIAEPVRLLR GRDKTGNAEL 251  FAWNIWVAGS GLAVIFWFDG AMAELLKFAM IAAFVSAPVF AWLNYRLVKG 301  DKRHRLTAGM NALAIVGLLY LAGFAVLFLL NLTGLLA*

ORF53ng-1 and ORF53-1 show 94.0% identity in 336 aa overlap:

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 58

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 487>:

  1 ..TTGCGGGAAA CGGCATATGT TTTGGATAGT TTTGATCGTT ATTTTGTTGT  51  TGCGCTTGCC GGCTTGTTTT TTGTCCGCGC ACAATCCGAA CGCGAGTGGA 101  TGCGCGAGGT TTCTGCGTGG CAGGAAAAGA AAGGGGAAAA ACAGGCGGAG 151  CTGCCTGAAA TCAAAGACGG TATGCCCGAT TTTCCCGAAC TTGCCCTGAT 201  GCTTTTCCAC GCCGTCAAAA CGGCAGTGTA TTGGCTGTTT GTCGGTGTCG 251  TCCGTTTCTG CCGAAACTAT CTGGCGCACG AATCCGAACC GGACAGGCCC 301  GTTCCGCCT..

This corresponds to the amino acid sequence <SEQ ID 488; ORF58>:

  1 ..LRETAYVLDS FDRYFVVALA GLFFVRAQSE REWMREVSAW QEKKGEKQAE  51  LPEIKDGMPD FPELALMLFH AVKTAVYWLF VGVVRFCRNY LAHESEPDRP 101   VPP..

Further work revealed the complete nucleotide sequence <SEQ ID 489>:

1 ATGTTTTGGA TAGTTTTGAT CGTTATTTTG TTGCTTGCGC TTGCCGGCTT 51 GTTTTTTGTCCGCGCACAAT CCGAACGCGA GTGGATGCGC GAGGTTTCTG 101 CGTGGCAGGA AAAGAAAGGGGAAAAACAGG CGGAGCTGCC TGAAATCAAA 151 GACGGTATGC CCGATTTTCC CGAACTTGCCCTGATGCTTT TCCATGCCGT 201 CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGTTTCTGCCGAA 251 ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC GCCTGCTTCT301 GCAAACCGTG CGGATGTTCC GACCGCATCC GACGGATATT CAGACAGTGG 351AAACGGGACG GAAGAAGCGG AAACGGAAGA AGCAGAAGCT GCGGAGGAAG 401 AGGCTGCCGATACGGAAGAC ATTGCAACTG CCGTAATCGA CAACCGCCGC 451 ATCCCATTCG ACCGGAGTATTGCTGAAGGG TTGATGCCGT CTGAAAGCGA 501 AATTTCGCCC GTCCGTCCGG TTTTTAAAGAAATCACTTTG GAAGAAGCAA 551 CGCGTGCTTT AAACAGCGCG GCTTTAAGGG AAACGAAAAAACGCTATATC 601 GATGCATTTG AGAAAAACGA AACAGCGGTC CCCAAAGTCC GCGTGTCCGA651 TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC CCTGTGCTTC 701AACGCACGTA TTCCCATATG TTCGATGCGG ACAAAGAAGC GTTTTCCGAG 751 TCTGCGGATTACGGATTTGA GCCGTATTTT GAGAAGCAGC ATCCGTCTGC 801 CTTTTCTGCA GTCAAAGCCGAAAATGCACG GAATGCGCCG TTCCACCGTC 851 ATGCAGGGCA GGGGAAAGGG CAGGCGGAGGCAAAATCCCC GGATGTTTCC 901 CAAGGGCAGT CCGTTTCAGA CGGCACGGCC GTCCGCGATGCCCGCCGCCG 951 CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT TCTGCGGAGG1001 CGCGAATTTC TCGCCTGATT CCGGAAAGTC AGACGGTTGT CGGGAAACGG 1051GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG AAACCGTTTC 1101 GTCTGTGGGATACGGCGGTC CGGTTTATGA TGAAACTGCC GATATCCATA 1151 TTGAAGAACC TGCCGCGCCCGATGCTTGGG TGGTCGAACC ACCCGAAGTG 1201 CCGAAAGTTC CCATGACCGC AATCGATATTCAGCCGCCGC CTCCCGTATC 1251 GGAAATCTAC AACCGTACCT ATGAACCGCC GTCAGGATTCGAGCAGGTGC 1301 AACGCAGCCG CATTGCCGAG ACCGACCATC TTGCCGATGA TGTTTTGAAT1351 GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCGGATGACG GCAGTGAAGG 1401TGCGGCAGAG CGGTCAAGCG GGCAATATCT GTCGGAAACC GAAGCGTTCG 1451 GGCATGACAGTCAGGCGGTT TGTCCGTTTG AAAATGTGCC GTCTGAACGC 1501 CCGTCCTGCC GGGTATCGGATACGGAAGCG GATGAAGGGG CGTTCCCATC 1551 TGAAGAAACC GGTGCGGTAT CCGAACACCTGCCGACAACC GACCTGCTTC 1601 TGCCTCCGCT GTTCAATCCC GAGGCGACGC AAACCGAAGAAGAACTGTTG 1651 GAAAACAGCA TCACCATCGA AGAAAAATTG GCGGAGTTCA AAGTCAAGGT1701 CAAGGTTGTC GATTCTTATT CCGGCCCCGT AATTACGCGT TATGAAATCG 1751AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTGAATCT GGAAAAAGAT 1801 TTGGCGCGTTCGCTCGGCGT GGCTTCCATC CGCGTTGTCG AAACCATCCC 1851 CGGCAAAACC TGCATGGGTTTGGAACTTCC GAACCCGAAA CGCCAAATGA 1901 TACGCCTGAG CGAAATCTTC AATTCGCCCGAGTTTGCCGA ATCCAAATCC 1951 AAGCTGACGC TCGCGCTCGG TCAGGACATC ACCGGACAGCCCGTCGTAAC 2001 CGACTTGGGA AAAGCACCGC ATTTGTTGGT TGCCGGCACG ACCGGTTCGG2051 GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT TTTCAAAGCC 2101GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA TGCTGGAATT 2151 GAGCATTTACGAAGGCATCC CGCACCTGCT CGCCCCTGTC GTTACCGATA 2201 TGAAGCTGGC GGCAAACGCGCTGAACTGGT GTGTTAACGA AATGGAAAAA 2251 CGCTACCGCC TGATGAGCTT TATGGGCGTGCGTAATCTTG CGGGCTTCAA 2301 TCAAAAAATC GCCGAAGCCG CAGCAAGGGG AGAAAAAATCGGCAATCCGT 2351 TCAGCCTCAC GCCCGACGAT CCCGAACCTT TGGAAAAACT GCCGTTTATC2401 GTGGTCGTGG TCGATGAGTT TGCCGACCTG ATGATGACGG CAGGCAAGAA 2451AATCGAAGAA CTGATTGCCC GCCTCGCCCA AAAAGCCCGC GCGGCAGGCA 2501 TCCATTTGATTCTTGCCACA CAACGCCCCA GCGTCGATGT CATCACGGGT 2551 CTGATTAAGG CGAACATCCCGACGCGTATC GCGTTCCAAG TGTCCAGCAA 2601 AATCGACAGC CGCACGATTC TCGACCAAATGGGCGCGGAA AACCTGCTCG 2651 GTCAGGGCGA TATGCTGTTC CTGCTGCCGG GTACTGCCTATCCGCAGCGC 2701 GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG TGGTCGAATA2751 TTTGAAACAG TTTGGCGAAC CGGACTATGT TGACGATATT TTGAGCGGCG 2801GCGGCAGCGA AGAGCTGCCC GGCATCGGGC GCAGCGGCGA CGACGAAACC 2851 GATCCGATGTACGACGAGGC CGTATCCGTT GTCCTGAAAA CGCGCAAAGC 2901 CAGCATTTCG GGCGTACAGCGCGCCTTGCG TATCGGCTAC AACCGCGCCG 2951 CGCGTCTGAT TGACCAGATG GAGGCGGAAGGCATTGTGTC CGCACCGGAA 3001 CACAACGGCA ACCGTACGAT TCTCGTCCCC TTGGACAATGCTTGA

This corresponds to the amino acid sequence <SEQ ID 490; ORF58-1>:

1 MFWIVLIVIL LLALAGL FFV RAQSEREWMR EVSAWQEKKG EKQAELPEIK 51 DGMPDFPELALMLFHAVKTA VYWLFVGVVR FCRNYLAHES EPDRPVPPAS 101 ANRADVPTAS DGYSDSGNGTEEAETEEAEA AEEEAADTED IATAVIDNRR 151 IPFDRSIAEG LMPSESEISP VRPVFKEITLEEATRALNSA ALRETKKRYI 201 DAFEKNETAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSHMFDADKEAFSE 251 SADYGFEPYF EKQHPSAFSA VKAENARNAP FHRHAGQGKG QAEAKSPDVS301 QGQSVSDGTA VRDARRRVSV NLKEPNKATV SAEARISRLI PESQTVVGKR 351DVEMPSETEN VFTETVSSVG YGGPVYDETA DIHIEEPAAP DAWVVEPPEV 401 PKVPMTAIDIQPPPPVSEIY NRTYEPPSGF EQVQRSRIAE TDHLADDVLN 451 GGWQEETAAI ADDGSEGAAERSSGQYLSET EAFGHDSQAV CPFENVPSER 501 PSCRVSDTEA DEGAFPSEET GAVSEHLPTTDLLLPPLFNP EATQTEEELL 551 ENSITIEEKL AEFKVKVKVV DSYSGPVITR YEIEPDVGVRGNSVLNLEKD 601 LARSLGVASI RVVETIPGKT CMGLELPNPK RQMIRLSEIF NSPEFAESKS651 KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN AMILSMLFKA 701APEDVRMIMI DPKMLELSIY EGIPHLLAPV VTDMKLAANA LNWCVNEMEK 751 RYRLMSFMGVRNLAGFNQKI AEAAARGEKI GNPFSLTPDD PEPLEKLPFI 801 VVVVDEFADL MMTAGKKIEELIARLAQKAR AAGIHLILAT QRPSVDVITG 851 LIKANIPTRI AFQVSSKIDS RTILDQMGAENLLGQGDMLF LLPGTAYPQR 901 VHGAFASDEE VHRVVEYLKQ FGEPDYVDDI LSGGGSEELPGIGRSGDDET 951 DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM EAEGIVSAPE1001 HNGNRTILVP LDNA*

Computer analysis of this amino acid sequence predicts the indicatedtransmembrane region, and also gave the following results:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF58 shows 96.6% identity over a 89aa overlap with an ORF (ORF58a) fromstrain A of N. meningitidis:

The complete length ORF58a nucleotide sequence <SEQ ID 491> is:

1 ATGTTTTGGA TAGTTTTGAT CGTTATTTTG TTGCTTGCGC TTGCCGGCTT 51 GTTTTTTGTCCGCGCACAAT CCGAACGCGA GTGGATGCGC GAGGTTTCTG 101 CGTGGCAGGA AAAGAAAGGGGAAAAACAGG CGGAGCTGCC TGAAATCAAA 151 GACGGTATGC CCGATTTTCC CGAACTTGCCCTGATGCTTT TCCATGCCGT 201 CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGTTTCTGCCGAA 251 ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC GCCTGCTTCT301 GCAAATCGTG CGGATGTTCC GACCGCATCC GACGGATATT CAGACAGTGG 351AAACGGGACG GAAGAAGCGG AAACGGAAGA AGCAGAAGCT GCGGAGGAAG 401 AGGCTGCCGATACGGAAGAC ATTGCAACTG CCGTAATCGA CAACCGCCGC 451 ATCCCATTCG ACCGGAGTATTGCTGAAGGG TTGATGCCGT CTGAAAGCGA 501 AATTTCGCCC GTCCGTCCGG TTTTTAAGGAAATCACTTTG GAAGAAGCAA 551 CGCGTGCTTT AAACAGCGCG GCTTTAAGGG AAACGAAAAAACGCTATATC 601 GATGCATTTG AGAAAAACGA AACAGCGGTC CCCAAAGTCC GCGTGTCCGA651 TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC CCTGTGCTTC 701AACGCACGTA TTCCCGTATG TTCGATGCGG ACAAAGAAGC GTTTTCCGAG 751 TCTGCGGATTACGGATTTGA GCCGTATTTT GAGAAGCAGC ATCCGTCTGC 801 CTTTTCTGCA GTCAAAGCCGAAAATGCACG GAATGCGCCG TTCCGCCGTC 851 ATGCAGGGCA GGGNAAAGGG CAGGCGGAGGCNAAATCCCC GGATGTTTCC 901 CAAGGGCAGT CCGTTTCAGA CGGCACAGCC GTCCGCGATGCCNGCCGCCG 951 CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT TCTGCGGAGG1001 CGCGGATTTC GCGCCTGATT CCGGAAAGTC GGACGGTTGT CGGGAAACGG 1051GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG AAANTGTTTC 1101 GTCTGTGGGATACGGCGNTC CGGTTTATGA TGAAACTGCC GATATCCATA 1151 TTGAAGAACC TGCCGCGCCCGATGCTTGGG TGGTCGAACC ACCCGAAGTG 1201 CCGAAAGTTC CCATGCCCGC AATNGATATTCCGCCGCCGC CTCCCGTATC 1251 GGAAATCTAC AACCGTACCT ATGAACCGCC GGCAGGATTCGAGCAGGTGC 1301 AACGCAGCCG CATTGCCGAA ACCGATCATC TTGCCGATGA TGTTTTGAAT1351 GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCGAATGACG GCAGTGAGGG 1401TGTGGCAGAG CGGTCAAGCG GGCAATATTT GTCGGAAACC GAAGCGTTCG 1451 GGCATGACAGTCAGGCGGTT TGTCCGTTTG AAAATGTGCC GTCTGAACGC 1501 CCGTCCCGCC GGGCATNGGATACGGAAGCG GATGAAGGGG CGTTCCAATC 1551 TGAAGAAACC GGTGCGGTAT CCGAACACCTGCCGACAACC GACCTGCTTC 1601 TGCCGCCGCT GTTCAATCCC GGGGCGACGC AAACCGAAGAAGANCTGTTG 1651 GANAACAGCA TCACCATCGA AGAAAAATNG GCGGAGTTCA AAGTCAAGGT1701 CAAGGTTGTC GATTCTTATT CCGGCCCCGT GATTACGCGT TATGAAATCG 1751AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTAAATCT GGAAAAAGAN 1801 TTGGCGCGTTCGCTCGGCGT GGCTTCCATC CGCGTTGTCG AAACCATCCT 1851 CGGCAAAACC TGTATGGGTTTGGAACTTCC GAACCCGAAA CGCCAAATGA 1901 TACGCCTGAG CGAAATCTTC AATTCGCCCGAGTTTGCCGA ATCCAAATCC 1951 AAGCTGACGC TCGCGCTCGG TCAGGACATC ACCGGACAGCCCGTCGTAAC 2001 CGACTTGGGC AAAGCACCGC ATTTGTTGGT TGCCGGCACG ACCGGTTCGG2051 GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT TTTCAAAGCC 2101GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA TGCTGGAATT 2151 GAGCATTTACGAAGGCATCC CGCACCTGCT CGCCCCTGTC GTTACCGATA 2201 TGAAGCTGGC GGCAAACGCGCTGAACTGGT GTGTTAACGA AATGGAAAAA 2251 CGCTACCGCC TGATGAGCTT TATGGGCGTGCGCAATCTTG CGGGTNTCAA 2301 TCAAAAAATC GCCGAAGCCG CAGCAAGGGG GGAGAAAATCGGCAACCCGT 2351 TCAGCCTCAC GCCCGACAAT CCCGAACCTT TGGANAAATT GCCGTTTATC2401 GTGGTCGTGG TTGATGAGTT TGCCGACCTG ATGATGACGG CAGGCAAGAA 2451AATCGAAGAA CTGATTGCCC GCCTCGCCCA AAAAGCCCGC GCGGCAGGCA 2501 TCCATCTTATCCTTGCCACA CAACGCCCCA GTGTCGATGT CATCACGGGT 2551 CTGATTAAGG CGAACATCCCGACGCGTATC GCGTTCCAAG TGTCCAGCAA 2601 AATCGACAGC CGCACGATTC TTGACCAAATGGGTGCGGAA AACCTGCTCG 2651 GGCAGGGCGA TATGCTGTTC CTGCCGCCGG GTACGGCCTATCCGCAGCGC 2701 GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG TGGTCGAATA2751 TCTGAAACAG TTTGGCGAAC CGGACTATGT TGACGATATN TTGAGCGGCG 2801GTATGTCCGA CGATTTGCTG GGAATCAGCC GGAGCGGCGA CGGCGAAACC 2851 GATCCGATGTACGACGAGGC CGTGTCNGTT GTTTTGAAAA CGCGCAAAGC 2901 CAGCATTTCT GGCGTGCAGCGCGCATTGCG TATCGGCTAT AATCGCGCCG 2951 CGCGTCTGAT TGACCAGATG GAGGCGGAAGGCATTGTGTC CGCACCGGAA 3001 CACAACGGCA ACCGTACGAT TCTCGTCCCC TTNGACAATGCTTGA

This encodes a protein having amino acid sequence <SEQ ID 492>:

1 MFWIVLIVIL LLALAGLFFV RAQSEREWMR EVSAWQEKKG EKQAELPEIK 51 DGMPDFPELALMLFHAVKTA VYWLFVGVVR FCRNYLAHES EPDRPVPPAS 101 ANRADVPTAS DGYSDSGNGTEEAETEEAEA AEEEAADTED IATAVIDNRR 151 IPFDRSIAEG LMPSESEISP VRPVFKEITLEEATRALNSA ALRETKKRYI 201 DAFEKNETAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSRMFDADKEAFSE 251 SADYGFEPYF EKQHPSAFSA VKAENARNAP FRRHAGQGKG QAEAKSPDVS301 QGQSVSDGTA VRDAXRRVSV NLKEPNKATV SAEARISRLI PESRTVVGKR 351DVEMPSETEN VFTEXVSSVG YGXPVYDETA DIHIEEPAAP wDAWVVEPPEV 401 PKVPMPAXDIPPPPPVSEIY NRTYEPPAGF EQVQRSRIAE TDHLADDVLN 451 GGWQEETAAI ANDGSEGVAERSSGQYLSET EAFGHDSQAV CPFENVPSER 501 PSRRAXDTEA DEGAFQSEET GAVSEHLPTTDLLLPPLFNP GATQTEEXLL 551 XNSITIEEKX AEFKVKVKVV DSYSGPVITR YEIEPDVGVRGNSVLNLEKX 601 LARSLGVASI RVVETILGKT CMGLELPNPK RQMIRLSEIF NSPEFAESKS651 KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN AMILSMLFKA 701APEDVRMIMI DPKMLELSIY EGIPHLLAPV VTDMKLAANA LNWCVNEMEK 751 RYRLMSFMGVRNLAGXNQKI AEAAARGEKI GNPFSLTPDN PEPLXKLPFI 801 VVVVDEFADL MMTAGKKIEELIARLAQKAR AAGIHLILAT QRPSVDVITG 851 LIKANIPTRI AFQVSSKIDS RTILDQMGAENLLGQGDMLF LPPGTAYPQR 901 VHGAFASDEE VHRVVEYLKQ FGEPDYVDDX LSGGMSDDLLGISRSGDGET 951 DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM EAEGIVSAPE1001 HNGNRTILVP XDNA*

ORF58a and ORF58-1 show 96.6% identity in 1014 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF58 shows complete identity over a 9aa overlap with a predicted ORF(ORF58ng) from N. gonorrhoeae:

The ORF58ng nucleotide sequence <SEQ ID 493> is predicted to encode aprotein having partial amino acid sequence <SEQ ID 494>:

1 ..SEPDRPVPPA SANRADVPTA SDGYSDSGNG TEEAETEAAE AAEEEAADTE 51  DIATAVIDNR RIPFDRSIAE GLMQSESKTS PVRPVFKEIT LEEATRALSS 101  AALRETKKRY IDAFEKNGTA VPKVRVSDTP MEGLQIIGLD DPVLQRTYSR 151  MFDADKEAFS ESADYGFEPY FEKQHPSAFS AVKAENARNA PFRRHAGQEK 201  GQAEAKSPDV SQGQSVSDGT AVRDARRRVS VNLKEPNKAT VSAEARISRL 251  IPESRTVVGK RDVEMPSETE NVFTETVSSV GYGGPVYDEA ADIHIEEPAA 301  PDAWVVEPPE VPEVAVPEID ILPPPPVSEI YNRTYEPPAG FEQAQRSRIA 351  ETDHLAADVL NGGWQEETAA IADDGSEGAA ERSSGQYLSE TEAFGHDSQA 401  VCPFEDVPSE RPSCRVSDTE ADEGAFQSEE TGAVSEHLPT TDLLLPPLFN 451  PEATQTEEEL LENSITIEEK LAEFKVKVKV VDSYSGPVIT RYEIEPDVGV 501  RGNSVLNLEK DLARSLGVAS IRVVETIPGK TCMGLELPNP KRQMIRLSEI 551  FNSPEFAESK SKLTLALGQD ITGQPVVTDL GKAPHLLVAG TTGSGKSVGV 601  NAMILSMLFK AAPEDVRMIM IDPKMLELSI YEGITHLLAP VVTDMKLAAN 651  ALNWCVNEME KRYRLMSFMG VRNLAGFNQK IAEAAARGEK IGNPFSLTPD 701  DPEPLEKLPF IVVVVDEFAD LMMTAGKKIE ELIARLAQKA RAAGIHLILA 751  TQRPSVDVIT GLIKANIPTR IAFQVSSKID SRTILDQMGA ENLLGQGDML 801  FLPPGTAYPQ RVHGAFASDE EVHRVVEYLK QFGEPDYVDD ILSGGGSEEL 851  PGIGRSGDGE TDPMYDEAVS VVLKTRKASI SGVQRALRIG YNRAARLIDQ 901  MEAEGIVSAP EHNGNRTILV PLDNA*

This partial gonococcal sequence contains a predicted transmembraneregion and a predicted ATP/GTP-binding site motif A (P-loop; doubleunderlined). Furthermore, it has a domain homologous to the FTSK celldivision protein of E. coli. Alignment of ORF58ng and FtsK (accessionnumber p46889) show a 65% amino acid identity in 459 overlap:

ORF58ng:  467 IEEKLAEFKVKVKVVDSYSGPVITRYEIEPDVGVRGNSVLNLEKDLARSLGVASIRVVET  526              +E +LA+F++K  VV+   GPVITR+E+    GV+   +NL +DLARSL    ++RVVEFtsK:     868 VEARLADFRIKADVVNYSPGPVITRFELNLAPGVKAARISNLSRDLARSLSTVAVRVVEV  927ORF58ng:  527 IPGKTCMGLELPNPKRQMIRLSEIFNSPEFAESKSKLTLALGQDITGQPVVTDLGKAPHL  586              IPGK  +GLELPN KRQ + L E+ ++ +F ++ S LT+LG+DI G+PVV DL K  PHLFtSK:     928 IPGKPYVGLELPNKKRQTVYLREVLDNAKFRDNPSPLTVVLGKDIAGEPVVADLAKMPHL  987ORF58ng:  587 LVAGTTGSGKSVGVNAMILSMLFKAAPEDVRMIMIDPKMLELSIYEGITHLLAPVVTDMK  646              LVAGTTGSGKSVGVNAMILSML+KA PEDVR IMIDPKMLELS+YEGI HLL  VVTDMKFtsK:     988 LVAGTTGSGKSVGVNAMILSMLYKAQPEDVRFIMIDPKMLELSVYEGIPHLLTEVVTDMK 1047ORF58ng:  647 LAANALNWCVNEMEKRYRLMSFMGVANLAGFNQKIAEAAARGEKIGNPFSLTPDDPEP--  704               AANAL WCVNEME+RY+LMS +GVRNLAG+N+KIAEA      I +P+     D  +FtsK:    1048 DAANALRWCVNEMERRYKLMSALGVRNLAGYNEKIAEADRMMRPIPDPYWKPGDSMDAQH 1107ORF58ng:  705 --LEKLPFIVVVVDEFADLMMTAGKKIEELIARLAQKARAAGIHLILATQRPSVDVITGL  762                L+K P+IVV+VDEFADLMMT GKK+EELIARLAQKARAAGIHL+LATQRPSVDVITGLFtsK:    1108 PVLKKEPYIVVLVDEFADLMMTVGKKVEELIARLAQKARAAGIHLVLATQRPSVDVITGL 1167ORF58ng:  763 IKANIPTRIAFQVSSKIDSRTILDQMGAENLLGQGDMLFLPPGTAYPQRVHGAFASDEEV  822              IKANIPTRIAF VSSKIDSRTILDQ GAE+LLG GDML+  P + P RVHGAF  D+EVFtsK:    1168 IKANIPTRIAFTVSSKIDSRTILDQAGAESLLGMGDMLYSGPNSTLPVRVHGAFVRDQEV 1227ORF58ng:  823 HRVVEYLKQFGEPDYVDDILSGGGSEELPGIGRSGDGETDPMYDEAVSVVLKTRKASISG   882              H VV+  K  G P YVD I S   SE   G G  G  E DP++D+AV  V +RKASISGFtsK:    1228 HAVVQDWKARGRPQYVDGITSDSESEGGAG-GFDGAEELDPLFDQAVQFVTEKRKASISG  1286ORF58ng:  883 VQRALRIGYNRAARLIDQMEAEGIVSAPEHNGNRTILVP                       921              VQR  RIGYNRAAR+I+QMEA+GIVS   HNGNR +L PFtsK:    1287 VQRQFRIGYNRAARIIEQMEAQGIVSEQGHNGNREVLAP                      1325

Further work on ORF58ng revealed the complete gonococcal DNA sequence tobe <SEQ ID 495>:

1 ATGTTTTGGA TAGTTTTGAT CGTTATtgtg TTGCTTGCGC TTGCCGGCCT 51 GTTTTTTGTCCGCGCACAAT CCGAACGCGA GTGGATGCGC GAGGTTTCTG 101 CGTGGCAGGA AAAGAAAGGGGAAAAACAGG CGGAGCTGCC TGAAATCAAA 151 GACGGTATGC CCGATTTTCC CGAGTTTTCCCTGATGCTTT TCCATGCCGT 201 CAAAACGGCA GTGTATTGGC TGTTTGTCGG TGTCGTCCGTTTCTGCCGAA 251 ACTATCTGGC GCACGAATCC GAACCGGACA GGCCCGTTCC GCCTGCTTCT301 GCAAACCGTG CGGATGTTCC GACCGCATCC GACGGGTATT CAGACAGTGG 351AAACGGGACG GAAGAAGCGG AAACGGAAGC AGCAGAAGCT GCGGAGGAAG 401 AGGCTGCCgATACgGAAGAC ATTGCAACTG CCGTAATCGA CAACCGCCGC 451 ATCCcatTCG ACCGGAGTATTGCTGAAGGG TTGATGCAGT CTGAAAGCAA 501 AACTTCGCCC GTCCGTCCGG TTTTTAAGGAAATCACTTTG GAAGAAGCAA 551 CGCGTGCTTT AAGCAGCGCG GCTTTAAGGG AAACGAAAAAACGCTATATC 601 GATGCATTTG AGAAAAACGG AACAGCCGTC CCCAAAGTAC GCGTGTCCGA651 TACCCCGATG GAAGGGCTGC AGATTATCGG TTTGGACGAC CCTGTGCTTC 701AACGCACGTA TTCCCGTATG TTTGATGCGG ACAAAGAAGC GTTTTCCGAG 751 TCTGCGGATTACGGATTTGA GCCGTATTTT GAGAAGCAGC ATCCGTCTGC 801 CTTTTCTGCA GTCAAAGCCGAAAATGCACG GAATGCGCCG TTCCGCCGTC 851 ATGCAGGGCA GGAGAAAGGG CAGGCGGAGGCAAAATCCCC GGATGTTTCC 901 CAAGGGCAGT CCGTTTCAGA CGGCACAGCC GTCCGCGATGCCCGCCGCCG 951 CGTTTCCGTC AATTTGAAAG AACCGAACAA GGCAACGGTT TCTGCGGAGG1001 CGCGGATTTC GCGCCTGATT CCGGAAAGTC GGACGGTTGT CGGGAAACGG 1051GATGTCGAAA TGCCGTCTGA AACCGAAAAT GTTTTCACGG AAACCGTTTC 1101 GTCTGTGGGATACGGCGGTC CGGTTTATGA TGAAGCTGCC GATATCCATA 1151 TTGAAGAGCC TGCCGCGCCCGATGCTTGGG TGGTCGAACC ACCCGAAGTG 1201 CCGGAGGTAG CCGTACCCGA AATCGATATTCTGCCGCCGC CTCCCGTATC 1251 GGAAATCTAC AACCGTACCT ATGAGCCGCC GGCAGGATTCGAGCAGGCGC 1301 AACGCAGCCG CATTGCCGAA ACCGACCATC TTGCCGCTGA TGTTTTGAAT1351 GGAGGTTGGC AGGAGGAAAC CGCCGCTATT GCAGATGACG GCAGTGAGGG 1401TGCGGCAGAG CGGTCAAGCG GGCAATATCT GTCGGAAACC GAAGCGTTCG 1451 GGCATGACAGTCAGGCGGTT TGTCCGTTTG AAGATGTGCC GTCTGAACGC 1501 CCGTCCTGCC GGGTATCGGATACGGAAGCG GATGAAGGGG CGTTCCAATC 1551 GGAAGAGACC GGTGCGGTAT CCGAACACCTGCCGACAACC GACCTGCTTC 1601 TGCCTCCGCT GTTCAATCCC GAGGCGACGC AAACCGAAGAAGAACTGTTG 1651 GAAAACAGCA TCACCATCGA AGAAAAATTG GCGGAGTTCA AAGTCAAGGT1701 CAAGGTTGTC GATTCTTATT CCGGCCCCGT GATTACGCGT TATGAAATCG 1751AACCCGATGT CGGCGTGCGC GGCAATTCCG TTCTGAATTT GGAAAAAGAC 1801 TTGGCGCGTTCGCTCGGCGT GGCTTCCATC CGCGTTGTCG AAACCATCCC 1851 CGGCAAAACC TGCATGGGTTTGGAACTTCC GAACCCGAAA CGCCAAATGA 1901 TACGCCTGAG CGAAATTTTC AATTCGCCCGAGTTTGCCGA ATCCAAATCC 1951 AAGCTGACGC TCGCGCTCGG TCAGGACATT ACCGGACAGCCCGTCGTAAC 2001 CGACTTGGGC AAAGCACCGC ATTTGCTGGT TGCCGGCACG ACCGGTTCGG2051 GCAAATCGGT GGGTGTCAAC GCGATGATTC TGTCTATGCT TTTCAAAGCC 2101GCGCCGGAAG ACGTGCGTAT GATTATGATC GATCCGAAAA TGCTGGAATT 2151 GAGCATTTACGAAGGCATCA CGCACCTGCT CGCCCCTGTC GTTACCGATA 2201 TGAAGCTGGC GGCAAACGCGCTGAACTGGT GTGTTAACGA AATGGAAAAA 2251 CGCTACCGCC TGATGAGCTT TATGGGCGTGCGCAATCTTG CGGGCTTCAA 2301 CCAAAAAATC GCCGAAGCCG CAGCAAGGGG AGAAAAAATCGGCAATCCGT 2351 TCAGCCTCAC GCCCGACGAT CCCGAACCTT TGGAAAAACT GCCGTTTATC2401 GTGGTCGTGG TCGATGAGTT TGCCGATTTG ATGATGACGG CAGGCAAGAA 2451AATCGAAGAA CTGATTGCGC GCCTCGCCCA AAAAGCCCGC GCGGCAGGCA 2501 TCCACCTTATCCTTGCCACA CAACGCCCCA GCGTCGATGT CATCACGGGT 2551 CTGATTAAGG CGAACATCCCGACGCGTATC GCGTTCCAAG TGTCCAGCAA 2601 AATCGACAGC CGCACGATTC TCGACCAAATGGGCGCGGAA AACCTGCTCG 2651 GTCAGGGCGA TATGCTGTTC CTGCCGCCGG GTACTGCCTATCCGCAGCGC 2701 GTTCACGGCG CGTTTGCCTC GGATGAAGAG GTGCACCGCG TGGTCGAATA2751 TCTGAAGCAG TTTGGCGAGC CGGACTATGT TGACGATATT TTGAGCGGCG 2801GCGGCAGCGA AGAGCTGCCC GGCATCGGGC GCAGCGGCGA CGGCGAAACC 2851 GATCCGATGTACGACGAGGC CGTATCCGTT GTCCTGAAAA CGCGCAAAGC 2901 CAGCATTTCG GGCGTACAGCGCGCCTTGCG CATCGGCTAC AACCGCGCCG 2951 CGCGTCTGAT TGACCAAATG GAAGCGGAAGGCATTGTGTC CGCACCGGAA 3001 CACAACGGCA ACCGTACGAT TCTCGTCCCC TTGGACAATGCTTGA

This corresponds to the amino acid sequence <SEQ ID 496; ORF58ng-1>:

1 MFWIVLIVIV LLALAGLFFV RAQSEREWMR EVSAWQEKKG EKQAELPEIK 51 DGMPDFPEFSLMLFHAVKTA VYWLFVGVVR FCRNYLAHES EPDRPVPPAS 101 ANRADVPTAS DGYSDSGNGTEEAETEAAEA AEEEAADTED IATAVIDNRR 151 IPFDRSIAEG LMQSESKTSP VRPVFKEITLEEATRALSSA ALRETKKRYI 201 DAFEKNGTAV PKVRVSDTPM EGLQIIGLDD PVLQRTYSRMFDADKEAFSE 251 SADYGFEPYF EKQHPSAFSA VKAENARNAP FRRHAGQEKG QAEAKSPDVS301 QGQSVSDGTA VRDARRRVSV NLKEPNKATV SAEARISRLI PESRTVVGKR 351DVEMPSETEN VFTETVSSVG YGGPVYDEAA DIHIEEPAAP DAWVVEPPEV 401 PEVAVPEIDILPPPPVSEIY NRTYEPPAGF EQAQRSRIAE TDHLAADVLN 451 GGWQEETAAI ADDGSEGAAERSSGQYLSET EAFGHDSQAV CPFEDVPSER 501 PSCRVSDTEA DEGAFQSEET GAVSEHLPTTDLLLPPLFNP EATQTEEELL 551 ENSITIEEKL AEFKVKVKVV DSYSGPVITR YEIEPDVGVRGNSVLNLEKD 601 LARSLGVASI RVVETIPGKT CMGLELPNPK RQMIRLSEIF NSPEFAESKS651 KLTLALGQDI TGQPVVTDLG KAPHLLVAGT TGSGKSVGVN AMILSMLFKA 701APEDVRMIMI DPKMLELSIY EGITHLLAPV VTDMKLAANA LNWCVNEMEK 751 RYRLMSFMGVRNLAGFNQKI AEAAARGEKI GNPFSLTPDD PEPLEKLPFI 801 VVVVDEFADL MMTAGKKIEELIARLAQKAR AAGIHLILAT QRPSVDVITG 851 LIKANIPTRI AFQVSSKIDS RTILDQMGAENLLGQGDMLF LPPGTAYPQR 901 VHGAFASDEE VHRVVEYLKQ FGEPDYVDDI LSGGGSEELPGIGRSGDGET 951 DPMYDEAVSV VLKTRKASIS GVQRALRIGY NRAARLIDQM EAEGIVSAPE1001 HNGNRTILVP LDNA*

ORF58ng-1 and ORF58-1 show 97.2% identity in 1014 aa overlap:

Furthermore, ORF58ng-1 shows significant homology to the E. coli proteinFtsK:

sp|P46889|FTSK_ECOLI CELL DIVISION PROTEINFTSK >gi|1651412|gnl|PID|d1015290 (Dl division protein FtsK [Escherichiacoli] >gi|1651418|gnl|PID|d1015296 (D90727) Cell division protein FtsK[Escherichia coli] >gi|1787117 (AE000191) cell division protein FtsK[Escherichia coli] Length = 1329 Score = 576 bits (1469), Expect = e−163Identities = 301/459 (65%), Positives = 353/459 (76%), Gaps = 5/459 (1%)Query: 556 IEEKLAEFKVKVKVVDSYSGPVITRYEIEPDVGVRGNSVLNLEKDLARSLGVASIRVVET615 +E +LA+F++K  VV+   GPVITR+E+    GV+   + NL +DLARSL   ++RVVE Sbjct:868 VEARLADFRIKADVVNYSPGPVITRFELNLAPGVKAARISNLSRDLARSLSTVAVRVVEV 927Query: 616 IPGKTCMGLELPNPKRQMIRLSEIFNSPEFAESKSKLTLALGQDITGQPVVTDLGKAPHL675 IPGK  +GLELPN KRQ + L E+ ++ +F ++ S LT+ LG+DI G+PVV DL K PHL Sbjct:928 IPGKPYVGLELPNKKRQTVYLREVLDNAKFRDNPSPLTVVLGKDIAGEPVVADLAKMPHL 987Query: 676 LVAGTTGSGKSVGVNAMILSMLFKAAPEDVRMIMIDPKMLELSIYEGITHLLAPVVTDMK735 LVAGTTGSGKSVGVNAMILSML+KA  PEDVR IMIDPKMLELS+YEGI HLL  VVTDMK Sbjct:988 LVAGTTGSGKSVGVNAMILSMLYKAQPEDVRFIMIDPKMLELSVYEGIPHLLTEVVTDMK 1047Query: 736 LAANALNWCVNEMEKRYRLMSFMGVRNLAGFNQKIAEAAARGEKIGNPFSLTPDDPEP--793  AANAL WCVNEME+RY+LMS +GVRNLAG+N+KIAEA      I +P+    D  + Sbjct:1048 DAANALRWCVNEMERRYKLMSALGVRNLAGYNEKIAEADRMMRPIPDPYWKPGDSMDAQH 1107Query: 794 --LEKLPFIVVVVDEFADLMMTAGKKIEELIARLAQKARAAGIHLILATQRPSVDVITGL851   L+K P+IVV+VDEFADLMMT GKK+EELIARLAQKARAAGIHL+LATQRPSVDVITGL Sbjct:1108 PVLKKEPYIVVLVDEFADLMMTVGKKVEELIARLAQKARAAGIHLVLATQRPSVDVITGL 1167Query: 852 IKANIPTRIAFQVSSKIDSRTILDQMGAENLLGQGDMLFLPPGTAYPQRVHGAFASDEEV911 IKANIPTRIAF VSSKIDSRTILDQ GAE+LLG GDML+  P +  P RVHGAF  D+EV Sbjct:1168 IKANIPTRIAFTVSSKIDSRTILDQAGAESLLGMGDMLYSGPNSTLPVRVHGAFVRDQEV 1227Query: 912 HRVVEYLKQFGEPDYVDDILSGGGSEELPGIGRSGDGETDPMYDEAVSVVLKTRKASISG971 H VV+  K  G P YVD I S   SE   G G  G  E DP++D+AV  V + RKASISG Sbjct:1228 HAVVQDWKARGRPQYVDGITSDSESEGGAG-GFDGAEELDPLFDQAVQFVTEKRKASISG 1286Query: 972 VQRALRIGYNRAARLIDQMEAEGIVSAPEHNGNRTILVP 1010VQR  RIGYNRAAR+I+QMEA+GIVS   HNGNR +L P Sbjct: 1287VQRQFRIGYNRAARIIEQMEAQGIVSEQGHNGNREVLAP 1325

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 59

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 497>:

This corresponds to the amino acid sequence <SEQ ID 498; ORF101>:

Further work revealed the complete nucleotide sequence <SEQ ID 499>:

1 ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA CCGCCGTCGG 51 CATTTTCGTCGTCCTCTTGG CGGTATTGGT CTCCACGCAG GCAATCAACC 101 TGCTCGGCCG TGCCGCCGACGGGCGTGTCG CCATCGATGC CGTGTTGGCA 151 TTGGTCGGCT TCTGGGTCAT CGGTATGACGCCGCTTTTGC TGGTGTTGAC 201 CGCATTTATC AGTACGTTGA CCGTGTTGAC CCGCTACTGGCGCGACAGCG 251 AAATGTCGGT CTGGCTATCC TGCGGATTGG CATTGAAACA ATGGATACGC301 CCGGTGATGC AGTTTGCCGT GCCGTTTGCC GTTTTGGTTG CCGTCATGCA 351GCTTTGGGTG ATACCGTGGG CAGAGCTACG CAGCCGCGAA TACGCTGAAA 401 TCCTGAAGCAGAAGCAGGAA TTGTCTTTGG TGGAGGCAGG CGAGTTCAAC 451 AGTTTGGGCA AGCGCAACGGCAGGGTTTAT TTTGTCGAAA CCTTCGATAC 501 CGAATCCGGC ATCATGAAAA ACCTGTTCCTGCGCGAACAG GACAAAAACG 551 GCGGCGACAA CATCATCTTC GCCAAAGAAG GTAACTTCTCGCTGAACGAC 601 AACAAACGCA CGCTCGAATT GCGCCACGGC TACCGTTACA GCGGCACGCC651 CGGACGCGCC GACTACAATC AGGTTTCCTT CCAAAAACTC AACCTGATTA 701TCAGCACCAC GCCCAAACTC ATCGACCCCG TTTCCCACCG CCGTACCATT 751 CCGACCGCCCAACTGATTGG CAGCAGCAAC CCGCAACATC AGGCGGAATT 801 GATGTGGCGC ATCTCGCTGACCGTCAGCGT CCTCCTACTC TGCCTGCTTG 851 CCGTGCCGCT TTCCTATTTC AACCCGCGCAGCGGACATAC CTACAATATC 901 TTGATTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGCTGACCCTGCT 951 TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC GGACTGCTGC1001 CTATGCACAT TATCATGTTT GCCGTTGCAC TCATCCTGTT GCGCGTCCGC 1051AGTATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA GTCTGACATT 1101 GAAAGGCGGAAAATGA

This corresponds to the amino acid sequence <SEQ ID 500; ORF101-1>:

1 MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD GRVAIDAVLA 51LVGFWVIGMT PLLLVLTAFI STLTVLTRYW RDSEMSVWLS CGLALKQWIR 101PVMQFAVPFA VLVAVMQLWV IPWAELRSRE YAEILKQKQE LSLVEAGEFN 151 SLGKRNGRVYFVETFDTESG IMKNLFLREQ DKNGGDNIIF AKEGNFSLND 201 NKRTLELRHG YRYSGTPGRADYNQVSFQKL NLIISTTPKL IDPVSHRRTI 251 PTAQLIGSSN PQHQAELMWRISLTVSVLLL CLLAVPLSYF NPRSGHTYNI 301 LIAIGLFLIY QNGLTLLFEA VEDGKIHFWLGLLPMHIIMF AVALILLRVR 351 SMPSQPFWQA VGKSLTLKGG K*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF101 shows 91.2% identity over a 57aa overlap and 95.7% identity overa 69aa overlap with an ORF (ORF101a) from strain A of N. meningitidis:

The complete length ORF101a nucleotide sequence <SEQ ID 501> is:

1 ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA CCGCCGTCGG 51 CATTTTCGTCGTCCTCTTGG CGGTATTGGT CTCCACGCAG GCAATCAACC 101 TGCTCGGCCN TGCCGCCGACNGGCGTNTCG CCATCGATGC CGTGTTGGCA 151 TTGGTCGGCT TCTGGGTCNN NNGNATGACGCCGCTTTTGC TNGTGTTGAC 201 CGCATTTATC AGTACGTTGA CCGTGTTGAC CCGCTACTGGCGNGACAGCG 251 AAATGTCGGT CTGGNTATCC TGCGGATTGG CATTGAAACA ATGGATACGC301 CCGGTGATGC AGTTTGCCGT GCCGTTTGCC GTTTTGGTTG CCGTCATGCA 351GCTTTGGGTG ATACCGTGGG CAGAGCTACG CAGCCGCGAA TACGCTGAAA 401 TCCTGAAGCAGAAGCAGGAA TTGTCTTTGG TGGAGGCAGG CGGGTTCAAC 451 AGTTTGGGCA AGCGCAACGGCAGGGTTTAT TTTGTCGAAA CCTTCGATAC 501 CGAATCCGGC ATCATGAAAA ACCTGTTCCTGCGCGAACAG GACAAAAACG 551 GCGGCGACAA CATCATCTTC NCCAAAGAAA GTAACTTCTCGCTGAACGAC 601 AACAAACGCA CGCTCGAATT GCGCCACGGC TACCGTTACA GCGGCACGCC651 CGGACGCGCC GACTACAATC AGGTTTCCTT CCNAAAACTC AACCTGATTA 701TCAGCACCAC GCCCAAACTC ATCGACCCCG TTTCCCACCG CCGTACNATN 751 CCNACNGCCCAACTGATTGG CAGCAGCAAC CCGCAACATC ANGCGGAATT 801 GATGTGGCGC ATCTCGCTGACCGTCAGCGT CCTCCTACTC TGCCTGCTTG 851 CCGTGCCGCT TTCCTATTTC AACCCGCGCAGCGGACATAC CTACAATATC 901 TTGANTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGCTGACCCTGCT 951 TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC GGACTGCTGC1001 CTATGCACAT CATCATGTTC GTCATCGCAA TCGTACTTCT GCGCGTCCGC 1051AGCATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA GTCTGACATT 1101 GAAAGGCGGAAAATGA

This encodes a protein having amino acid sequence <SEQ ID 502>:

1 MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGXAAD XRXAIDAVLA 51LVGFWVXXMT PLLLVLTAFI STLTVLTRYW RDSEMSVWXS CGLALKQWIR 101PVMQFAVPFA VLVAVMQLWV IPWAELRSRE YAEILKQKQE LSLVEAGGFN 151 SLGKRNGRVYFVETFDTESG IMKNLFLREQ DKNGGDNIIF XKESNFSLND 201 NKRTLELRHG YRYSGTPGRADYNQVSFXKL NLIISTTPKL IDPVSHRRTX 251 PTAQLIGSSN PQHXAELMWRISLTVSVLLL CLLAVPLSYF NPRSGHTYNI 301 LXAIGLFLIY QNGLTLLFEA VEDGKIHFWLGLLPMHIIMF VIAIVLLRVR 351 SMPSQPFWQA VGKSLTLKGG K*

ORF101a and ORF101-1 show 95.4% identity in 371 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF101 shows 96.5% identity in 57aa overlap at the N-terminal domain and95.1% identity in 61 as overlap at the C-terminal domain, respectively,with a predicted ORF (ORF101ng) from N. gonorrhoeae:

The ORF101ng nucleotide sequence <SEQ ID 503> is predicted to encode aprotein having partial amino acid sequence <SEQ ID 504>:

1 MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD GRVAIDAVLA 51 LVGFWVIGMTPLLLVLTAFI STLTVLTRYW RDSEMSVWLS CGLALKQWIR 101PVMQFAVPFA ILIAVMQLWV IPWAELRSRE YAEILKQKQE LSLVEAGEFN 151 NLGKRNGRVYFVETFDTESG IMKNLFLREQ DKNGGDNIIF AKEGNFSLKD 201 NKRTLELRHG YRYSGTPGRADYNQVSFQKL NLIISTTPKL IDPVSHRRTI 251 STAQLIGSSN PQHQAELMWRISLTVSVLLL CLLAVPLSYF NPRSGHTYNI 301 LIAIGLFLIY QNGLTLLFEA VEDGKIHFWLGLLPMHIIMF VIAIVLLRVR 351 SMPSQPFWQA VG...

Further work revealed the complete nucleotide sequence <SEQ ID 505>:

1 ATGATTTATC AAAGAAACCT CATCAAAGAA CTCTCTTTTA CCGCCGTCGG 51 CATTTTCGTCGTCCTCTTGG CGGTGTTGGT GTCCACGCAG GCGATCAACC 101 TGCTTGGCCG CGCAGCTGACGGGCGTGTCG CCATCGATGC CGTGTTGGCC 151 TTAGTCGGCT TCTGGGTCAT CGGTATGACCCCGCTTTTGC TGGTGTTGAC 201 CGCATTCATC AGCACGCTGA CCGTATTGAC CCGCTACTGGCGCGACAGCG 251 AAATGTCGGT CTGGCTATCC TGCGGATTGG CGTTGAAACA GTGGATACGC301 CCCGTCATGC AGTTTGCCGT GCCGTTTGCC ATCCTGATTG CCGTCATGCA 351GCTTTGGGTG ATACCGTGGG CAGAGCTGCG CAGCCGCGAA TATGCCGAAA 401 TTTTGAAGCAGAAGCAGGAA TTGTCTTTGG TGGAAGCCGG CGAGTTCAAT 451 AACTTGGGCA AGCGCAACGGCAgggtttaT TtcgtcgaaA CCTTTGACAC 501 CGaatccgGC ATCATGAAAA ACCTGTtcctGcGCGAACAG GACAAAAACG 551 gcggcgacaA CATCATCTTC GCcaaaGAag gtaactTctcgctgaaggaC 601 AACAAAcgca cgctcgaATT GCGCCACGGC TACCGTTACA GCGGcacgcC651 CGGacGCGCc gactaCAATC AGGTTtcctt cCAAAAacTc aacctgATta 701TCAGCACCAC GCCCAAacTT ATCGaccCCG TTTCCCACCG CCGCACCATT 751 tcgacCGCCCAAcTGATTGG CAGCAGCAAT CCGCAACATC AGGCAGAATT 801 GATGTGGCGC ATCTCGCTGACCGTCAGCGT CCTCCTGCTC TGCCTACTCG 851 CCGTGCCGCT TTCCTATTTC AACCCGCGCAGCGGACATAC CTACAATATC 901 TTGATTGCCA TCGGTTTGTT TTTAATTTAC CAAAACGGGCTGACCCTGCT 951 TTTTGAAGCC GTGGAAGACG GCAAAATCCA TTTTTGGCTC GGACTGCTGC1001 CTATGCACAT CATCATGTTC GTCATCGCAA TCGTACTTCT GCGCGTCCGC 1051AGTATGCCCA GCCAGCCCTT CTGGCAGGCG GTTGGCAAAA GTCTGACATT 1101 GAAAGgcgGAAAATGA

This corresponds to the amino acid sequence <SEQ ID 506; ORF101ng-1>:

1 MIYQRNLIKE LSFTAVGIFV VLLAVLVSTQ AINLLGRAAD GRVAIDAVLA 51LVGFWVIGMT PLLLVLTAFI STLTVLTRYW RDSEMSVWLS CGLALKQWIR 101PVMQFAVPFA ILIAVMQLWV IPWAELRSRE YAEILKQKQE LSLVEAGEFN 151 NLGKRNGRVYFVETFDTESG IMKNLFLREQ DKNGGDNIIF AKEGNFSLKD 201 NKRTLELRHG YRYSGTPGRADYNQVSFQKL NLIISTTPKL IDPVSHRRTI 251 STAQLIGSSN PQHQAELMWRISLTVSVLLL CLLAVPLSYF NPRSGHTYNI 301 LIAIGLFLIY QNGLTLLFEA VEDGKIHFWLGLLPMHIIMF VIAIVLLRVR 351 SMPSQPFWQA VGKSLTLKGG K*

ORF101ng-1 and ORF101-1 show 97.6% identity in 371 aa overlap:

Based on this analysis, including the presence of a putative leadersequence (double-underlined) and several putative transmembrane domains(single-underlined) in the gonococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 60

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 507>:

1 ..GGTGGTGGTT TTATCAATGC TTCCTGTGCC ACTTTGACGA CAGCCAAACC 51  GCAATATCAA GCAGGAGACC TTAGCGCTTT TAAGATAAGG CAAGGCAATG 101  TTGTAATCGC CGGACACGGT TTGGATGCAC GTGATACCGA TTACACACGT 151  ATTCTCAGTT ATCATTCCAA AATCGATGCA CCCGTATGGG GACAAGATGT 201  TCGTGTCGTC GCGGGACAAA ACGATGTGGC CGCAACAGGT GATGCACATT 251  CGCCTATTCT CAATAATGCT GCTGCCAATA CGTCAAACAA TACAGCCAAC 301  AACGGCACAC ATATCCCTTT ATTTGCGATT GATACAGGCA AATTAGGAGG 351  TAT.GTATGC CAACAAAATC ACCTTGATCA GTACGGTCGA GCAAGCAGGC 401   ATTCGTAA

This corresponds to the amino acid sequence <SEQ ID 508; ORF113>:

1 ..GGGFINASCA TLTTAKPQYQ AGDLSAFKIR QGNVVIAGHG LDARDTDYTR 51  ILSYHSKIDA PVWGQDVRVV AGQNDVAATG DAHSPILNNA AANTSNNTAN 101  NGTHIPLFAI DTGKLGGXVC QQNHLDQYGR ASRHS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with pspA Putative Secreted Protein of N. meningitidis(Accession AF030941)

ORF and pspA show 44% aa identity in 179aa overlap:

orf113 GGGFINASCATLTTAKPQYQAGDLSAFKIRQGNVVIAGHGLDARDTDYTRILSYHSKIDA 60GGG INA+  TLT+  P    G+L+ F +  G VVI G GLD  D DYTRILS  ++I+A pspaGGGLINAASVTLTSGVPVLNNGNLTGFDVSSGKVVIGGKGLDTSDADYTRILSRAAEINA 256 orf113PVWGQDVRVVAGQNDVAATGDAHSPILXXXXXXXXXXXXXXGTHIPLFAIDTGKLGGMYA 120 VWG+DV+VV+G+N +   G                      +  P  AIDT  LGGMYA pspaGVWGKDVKVVSGKNKLDFDG---------SLAKTASAPSSSDSVTPTVAIDTATLGGMYA 307 orf113NKITLISTVEQAGIRNQGQWFASAGNVAVNAEGKLVNTGMIAATGENHAVSLHARNVHN 179+KITLIST   A IRN+G+ FA+ G V ++A+GKL N+G I A      +++ A+ V N pspaDKITLISTDNGAVIRNKGRIFAATGGVTLSADGKLSNSGSIDAA----EITISAQTVDN 362Homology with a Predicted ORF from N. gonorrhoeae

ORF113 shows 86.5% identity in 52aa overlap at the N-terminal part and94.1% identity in 17aa overlap at the C-terminal part with a predictedORF (ORF113ng) from N. gonorrhoeae:

The complete length ORF113ng nucleotide sequence <SEQ ID 509> ispredicted to encode a protein having amino acid sequence <SEQ ID 510>:

1 MNKTLYRVIF NRKRGAVVAV AETTKREGKS CADSGSGSVY VKSVSFIPTH 51 SKAFCFSALGFSLCLALGTV NIAFADGIIT DKAAPKTQQA TILQTGNGIP 101 QVNIQTPTSA GVSVNQYAQFDVGNRGAILN NSRSNTQTQL GGWIQGNPWL 151 TRGEARVVVN QINSSHPSQL NGYIEVGGRRAEVVIANPAG IAVNGGGFIN 201 ASRATLTTGQ PQYQAGDFSG FKIRQGNAVI AGHGLDARDTDFTRILVCQQ 251 NHLDQYGRTS RHS*

Based on this analysis, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 61

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 511>:

1 ..TCAACGGGAC ATAGCGAACA AAATTACACT TTGCCGCGAG AAATCACACG 51  CAACATTTCA CTGGGTTCAT TTGCCTATGA ATCGCATCGC AAAGCATTAA 101  GCCATCATGC GCCCAGCCAA GGCACTGAGT TGCCGCAAAG CAACGGTATT 151  TCGCTACCCT ATACGTCCAA TTCTTTTACC CCATTACCCA GCAGCAGCTT 201  ATACATTATC AATCCTGTCA ATAAAGGCTA TCTTGTTGAA ACCGATCCAC 251  GCTTTGCCAA CTACCGTCAA TGGTTGGGTA GTGACTATAT GCtGGACAGC 301  CTCAAACTAG ACCCAAACAA TTTACATAAA CGTTTGGGTG ATGGTTATTA 351  CGAGCAACGT TTAATCAATG AACAAATCGC AGAGCTGACA GGGCATCGTC 401  GTTTAGAcGG TTATCAAAAC GACGAAGAAC AATTTAAAGC CTTAATGGAT 451  AATGGCGCGA CTGCGGCACG TTcGATGAAT CTCAGCGTTG GCATTGCATT 501  AAGTGCCGAG CAAGTAGCGC AACTGACCAG CGATATTGTT TGGTTGGTAC 551  AAAAAGAAGT TAAGCTTCCT GATGGCGGCA CACAAACCGT ATTGGTGCCA 601  CAGGTTTATG TACGCGTTAA AAATGGCGAC ATAGACGGTA AAGGTGCATT 651  GTTGTCAGGC AGCAATACAC AAATCAATGT TTCAGGCAGC CTGAAAAACT 701  CAGGCACGAT TGCAGGgCGC AATGCGCTTA TTATCAATAC CGATACGCTA 751  GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGG TTACGGCCAC 801  ACAAGACATC AATAATATTG GCGGCATGCT TTCTGCCGAA CAGACATTAT 851  TGCTCAACGC AGGCAACAAC ATCAACAGCC AAAGCACCAC CGCCAGCAGT 901  CAAAATACAC AAGGCAGCAG CACCTACCTA GACCGAATGG CAGGTATTTA 951  TATCACAGGC AAAGAAAAAG GTGTTT..

This corresponds to the amino acid sequence <SEQ ID 512; ORF115>:

1 ..STGHSEQNYT LPREITRNIS LGSFAYESHR KALSHHAPSQ GTELPQSNGI 51  SLPYTSNSFT PLPSSSLYII NPVNKGYLVE TDPRFANYRQ WLGSDYMLDS 101  LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN DEEQFKALMD 151  NGATAARSMN LSVGIALSAE QVAQLTSDIV WLVQKEVKLP DGGTQTVLVP 201  QVYVRVKNGD IDGKGALLSG SNTQINVSGS LKNSGTIAGR NALIINTDTL 251  DNIGGRIHAQ KSAVTATQDI NNIGGMLSAE QTLLLNAGNN INSQSTTASS 301  QNTQGSSTYL DRMAGIYITG KEKGV..

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the pspA Putative Secreted Protein of N. meningitidis(Accession Number AF030941)

ORF115 and pspA protein show 50% aa identity in 325aa overlap:

Orf115: 1 STGHSEQNYTLPREITRNISLGSFAYESHRKALSHHAPSQGTELPQSNGISLPYTSNSFT60 STG+S   Y    E++ +I +G  AY+ +    +   P     +   NGI   +T pspA: 778STGYSRSPYEPAPEVS-SIRMGISAYKGYAPQQASDIPGTVVPVVAENGIHPTFT----- 831 Orf115:61 PLPSSSLYIINPVNKGYLVETDPRFANYRQWLGSDYMLDSLKLDPNNLHKRLGDGYYEQR 120 LP+SSL+ I P NKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+ pspA: 832-LPNSSLFAIAPNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQK 890 Orf115:121 LINEQIAELTGHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQVAQLTSDIV 180L+NEQIA+LTG+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQVA+LTSDIV pspA: 891LVNEQIAKLTGYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIV 950 Orf115:181 WLVQKEVKLPDGGTQTVLVPQVYVRVKNGDIDGKGALLSGSNTQINVSGSLKN-SGTIAG 239WL  + V LPDG TQTVL P+VYVR +  D++G+GALLSGS   I  SG+++N  G IAG pspA: 951WLENETVTLPDGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAG 1009Orf115: 240 RNALIINTDTLDNIGGRIHAQKSAVTATQDINNIGGMLSAEQTLLLNAGXXXXXXXXXXX299 R ALI+N   + N+ G +  +     A  DI N G  + AE  LLL A pspA: 1010REALILNAQNIKNLQGDLQGKNIFAAAGSDITNTGS-IGAENALLLKASNNIESRSETRS 1068Orf115: 300 XXXXXXXXXYLDRMAGIYITGKEKG 324           + R+AGIY+TG++ GpspA: 1069 NQNEQGSVRNIGRVAGIYLTGRQNG 1093Homology with a Predicted ORF from N. gonorrhoeae

ORF115 shows 91.9% identity over a 334aa overlap with a predicted ORF(ORF115ng) from N. gonorrhoeae:

An ORF115ng nucleotide sequence <SEQ ID 513> was predicted to encode aprotein having amino acid sequence <SEQ ID 514>:

1 MLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD ETGHREQNYT 51 LPEEITRDISLGSFAYESHS KALSRHAPSQ GTELPQSNRD NIRTAKSNGI 101 SLPYTPNSFT PLPGSSLYIINPANKGYLVE TDPRFANYRQ WLGSDYMLGS 151 LKLDPNNLHK RLGDGYYEQR LINEQIAELTGHRRLDGYQN DEEQFKALMD 201 NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLPDGGTQTVLMP 251 QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR NALIINTDTL301 DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN INNQSTAKSS 351QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ ISNQSDQGQT 401 RLQAGRDINLDTVQTGKYQE IHFDADNHTI RGSTNEVGSS IQTKGDVTLL 451 SGNNLNAKAA EVGSAKGTLAVYAKNDITIS SGIHAGQVDD ASKHTGRSGG 501 GNKLVITDKA QSHHETAQSS TFEGKQVVLQAGNDANILGS NVISDNGTRI 551 QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSKTNTQENQSQS 601 NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS TQSMDIGAAQ651 NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAK QFDKAKTTAL 701MPWRLPMQVG RLFKQAKAPK K*

Further work revealed the following partial gonococcal DNA sequence <SEQID 515>:

1 TTGCTTGTGC AAACAGAAAA AGACGGTTTG CATAACGAGC AAACCTTTGG 51 CGAGAAGAAAGTCTTCAGCG AAAATGGTAA GTTGCACAAC TACTGGCGTG 101 CGCGTCGTAA AGGACATGATGAAACAGGGC ATCGTGAACA AAATTATACT 151 TTGCCGGAGG AAATCACACG CGACATTTCACTGGGTTCAT TTGCCTATGA 201 ATCGCATAGC AAAGCATTAA GCCGTCATGC GCCCAGCCAAGGCACTGAGT 251 TGCCACAAAG TAACCGGGAT AATATCCGTA CTGCGAAAAG CAACGGTATT301 TCGCTACCCT ATACGCCCAA TTCTTTTACC CCATTACCCG GCAGCAGCTT 351ATACATTATC AATCCTGCCA ATAAAGGCTA TCTTGTTGAA ACCGATCCAC 401 GCTTTGCCAACTACCGTCAA TGGTTGGGTA GTGACTATAT GCTGGGCAGC 451 CTCAAACTAG ACCCAAACAATTTACATAAA CGTTTGGGTG ATGGTTATTA 501 CGAGCAACGT TTAATCAATG AACAAATCGCAGAGCTGACA GGGCATCGTC 551 GTTTAGACGG TTATCAAAAC GACGAAGAAC AATTTAAAGCCTTAATGGAT 601 AATGGCGCGA CTGCGGCACG TTCGATGAAT CTCAGCGTTG GCATTGCATT651 AAGTGCCGAG CAAGCAGCGC AACTGACCAG CGATATTGTT TGGTTGGTAC 701AAAAAGAAGT TAAACTTCCT GATGGCGGCA CACAAACCGT ATTGATGCCA 751 CAGGTTTATGTACGCGTTAA AAATGGCGGC ATAGACGGTA AAGGTGCATT 801 GTTGTCAGGC AGCAATACACAAATCAATGT TTCAGGCAGC CTGAAAAACT 851 CAGGCACGAT TGCAGGGCGC AATGCGCTTATTATCAATAC CGATACGCTA 901 GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGGTTACGGCCAC 951 ACAAGACATC AATAATATTG GCGGCATTCT TTCTGCCGAA CAGACATTAT1001 TGCTCAATGC GGGTAACAAC ATCAACAACC AAAGCACGGC CAAGAGCAGT 1051CAAAATGCAC AAGGTAGCAG CACCTACCTA GACCGAATGG CAGGTATTTA 1101 TATCACAGGCAAAGAAAAAG GTGTTTTAGC AGCGCAGGCA GGCAAAGACA 1151 TCAACATCAT TGCCGGTCAAATCAGCAATC AATCAGATCA AGGGCAAACC 1201 CGGCTGCAGG CAGGACGCGA CATTAACCTGGATACGGTAC AAACCGGCAA 1251 ATATCAAGAA ATCCATTTTG ATGCCGATAA CCATACCATCCGAGGTTCAA 1301 CGAACGAAGT CGGCAGCAGC ATTCAAACAA AAGGCGATGT TACCCtatTG1351 TCAGGGAATA ATCTCAATGC CAAAGCTGCC GAAGTCGGCA GCGCAAAAGG 1401CACACTTGCC GTGTATGCTA AAAATGACAT TACTATCAGC TCAGGCATCC 1451 ATGCCGGCCAAGTTGATGAT GCGTCCAAAC ATACAGGCAG AAGCGGCGGC 1501 GGTAATAAAT TAGTCATTACCGATAAAGCC CAAAGTCATC ACGAAACTGC 1551 TCAAAGCAGC ACCTTTGAAG GCAAGCAAGTTGTATTGCAG GCAGGAAACG 1601 ATGCCAACAT CCTTGGCAGT AATGTTATTT CCGATAATGGCACCCGGATT 1651 CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC AAAGCCAAAG1701 CGAAACCTAT CATCAAACCC AAAAATCAGG ATTGATGAGT GCAGGTATCG 1751GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA ATCCCAAAGC 1801 AACGAACATACAGGCAGTAC CGTAGGCAGC CTGAAAGGCG ATACCACCAT 1851 TGTTGCAAGC AAACACTACGAACAAACCGG CAGCAACGTT TCCAGCCCTG 1901 AGGGCAACAA CCTTATCAGC ACGCAAAGTATGGATATTGG CGCAGCACAA 1951 AACCAATTAA ACAGCAAAAC CACCCAAACC TACGAACAAAAAGGCTTAAC 2001 GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA GCGATTGCCG2051 TAGCACACAA AGCAGCAAAC AAGTCGGACA AAGCAAAAAC GACCGCGTTA 2101ATGCCATGGC GGCTGCCAAT GCAGGTTGGC AGGCCTATCA AACAGGCAAA 2151 GGCGCACAAAACTTAG

This corresponds to the amino acid sequence <SEQ ID 516; ORF115ng-1>:

1 LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD ETGHREQNYT 51 LPEEITRDISLGSFAYESHS KALSRHAPSQ GTELPQSNRD NIRTAKSNGI 101 SLPYTPNSFT PLPGSSLYIINPANKGYLVE TDPRFANYRQ WLGSDYMLGS 151 LKLDPNNLHK RLGDGYYEQR LINEQIAELTGHRRLDGYQN DEEQFKALMD 201 NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLPDGGTQTVLMP 251 QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR NALIINTDTL301 DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN INNQSTAKSS 351QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ ISNQSDQGQT 401 RLQAGRDINLDTVQTGKYQE IHFDADNHTI RGSTNEVGSS IQTKGDVTLL 451 SGNNLNAKAA EVGSAKGTLAVYAKNDITIS SGIHAGQVDD ASKHTGRSGG 501 GNKLVITDKA QSHHETAQSS TFEGKQVVLQAGNDANILGS NVISDNGTRI 551 QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSKTNTQENQSQS 601 NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS TQSMDIGAAQ651 NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAN KSDKAKTTAL 701MPWRLPMQVG RPIKQAKAHK T*

This gonococcal protein (ORF115ng-1) shows 91.9% identity with ORF115over 334aa:

In addition, it shows homology with a secreted N. meningitidis proteinin the database:

gi|2623258 (AF030941) putative secreted protein [Neisseria meningitidis]Length = 2273 Score = 604 bits (1541), Expect = e−172 Identities= 325/678 (47%), Positives = 449/678 (65%), Gaps = 22/678 (3%) Query: 1LLVQTEKDGLHNEQTFGEKKVFSENGKLHNYWRARRKGHDETGHREQNYTLPEEITRDIS 60 L+V T+  L N++T G K + ++ G LH Y R  +KG D TG+    Y    E++  I Sbjct: 739LIVGTPESALDNDETLGTKTI-TDKGDLHRYHRHHKKGRDSTGYSRSPYEPAPEVS-SIR 796 Query:61 LGSFAYESHSKALSRHAPSQGTELPQSNRDNIRTAKSNGISLPYTPNSFTPLPGSSLYII 120+G  AY+ +       AP Q +++P +    +     NGI   +T      LP SSL+ I Sbjct: 797MGISAYKGY-------APQQASDIPGTV---VPVVAENGIHPTFT------LPNSSLFAI 840 Query:121 NPANKGYLVETDPRFANYRQWLGSDYMLGSLKLDPNNLHKRLGDGYYEQRLINEQIAELT 180  PNKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+L+NEQIA+LT Sbjct: 841APNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQKLVNEQIAKLT 900 Query:181 GHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQAAQLTSDIVWLVQKEVKLP 240G+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQ A+LTSDIVWL  + V LP Sbjct: 901GYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIVWLENETVTLP 960 Query:241 DGGTQTVLMPQVYVRVKNGGIDGKGALLSGSNTQINVSGSLKN-SGTIAGRNALIINTDT 299 DGTQTVL P+VYVR +   ++G+GALLSGS   I  SG+++N  G IAGR ALI+N Sbjct: 961DGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAGREALILNAQN 1019 Query:300 LDNIGGRIHAQKSAVTATQDINNIGGILSAEQTLLLNAGNNINNQSTAKSSQNAQGSSTY 359+ N+ G +  +     A  DI N G I  AE  LLL A NNI ++S  +S+QN QGS Sbjct: 1020IKNLQGDLQGKNIFAAAGSDITNTGSI-GAENALLLKASNNIESRSETRSNQNEQGSVRN 1078 Query:360 LDRMAGIYITGKEKGVLAAQAGKDINIIAGQISNQSDQGQTRLQAGRDINLDTVQTGKYQ 419+ R+AGIY+TG++ G +   AG +I + A +++NQS+ GQT L AG DI  DT    + Q Sbjct: 1079IGRVAGIYLTGRQNGSVLLDAGNNIVLTASELTNQSEDGQTVLNAGGDIRSDTTGISRNQ 1138 Query:420 EIHFDADNHTIRGSTNEVGSSIQTKGDVTLLSGNNLNAKAAEVGSAKGTLAVYAKNDITI 479   FD+DN+ IR   NEVGS+I+T+G+++L +  ++  +AAEVGS + G L + A  DI + Sbjct:1139 NTIFDSDNYVIRKEQNEVGSTIRTRGNLSLNAKGDIRIRAAEVGSEQGRLKLAAGRDIKV 1198Query: 480 SSGIHAGQVDDASKHTGRSGGGNKLVITDKAQSHHETAQSSTFEGKQVVLQAGNDANILG539  +G    + +DA K+TGRSGGG K  +T   ++ +  A S T +GK+++L +G D  + G Sbjct:1199 EAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQNGQAVSGTLDGKEIILVSGRDITVTG 1258Query: 540 SNVISDNGTRIQAGNHVRIGTTQTQSQSETYHQTQKSGLM-SAGIGFTIGSKTNTQENQS598 SN+I+DN T + A N++ +   +T+S+S   ++ +KSGLM S GIGFT GSK +TQ N+S Sbjct:1259 SNIIADNHTILSAKNNIVLKAAETRSRSAEMNKKEKSGLMGSGGIGFTAGSKKDTQTNRS 1318Query: 599 QSNEHTGSTVGSLKGDTTIVASKHYEQTGSNVSSPEGNNLISTQSMDIGAAQNQLNSKTT658 ++  HT S VGSL G+T I A KHY QTGS +SSP+G+  IS+  + I AAQN+ + ++ Sbjct:1319 ETVSHTESVVGSLNGNTLISAGKHYTQTGSTISSPQGDVGISSGKISIDAAQNRYSQESK 1378Query: 659 QTYEQKGLTVAFSSPVTD 676 Q YEQKG+TVA S PV + Sbjct: 1379QVYEQKGVTVAISVPVVN 1396

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 62

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 517>:

1 ..TCAGGGAATA ACCTCAATGC CAAAGCTGCC GAAGTCAGCA GCGCAAACGG 51  TACACTCGCT GTGTCTGCCA ATAATGACAT CAACATCAGC GCAGGCATCA 101  ACACGACCCA TGTTGATGAT GCGTCCAAAC ACACAGGCAG AAGCGGTGGT 151  GGCAATAAAT TAGTCATTAC CGATAAAGCC CAAAGTCATC ACGAAACCGC 201  CCAAAGCAGC ACCTTTGAAG GCAAGCAAGT TGTATTGCAG GCAGGAAACG 251  ATGCCAACAT CCTTGGCAGC AATGTTATTT CCGATAATGG CACCCAGATT 301  CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC AAAGCCAAAG 351  CGAAACCTAT CATCAAACCC AGAAATCAGG ATTGATGAGT GCAGGTATCG 401  GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA ATCCCAAAGC 451  AACGAACATA CAGGCAGTAC CGTAGGCAGC TTGAAAGGCG ATACCACCAT 501  TGTTGCAGGC AAACACTACG AACAAATCGG CAGTACCGTT TCCAGCCCGG 551  AAGGCAACAA TACCATCTAT GCCCAAAGCA TAGACATTCA AGCGGCACAC 601  AACAAATTAA ACAGTAATAC CACCCAAACC TATGAACAAA AAGG.CTAAC 651  GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA ...

This corresponds to the amino acid sequence <SEQ ID 518; ORF117>:

1 ..SGNNLNAKAA EVSSANGTLA VSANNDINIS AGINTTHVDD ASKHTGRSGG 51  GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS NVISDNGTQI 101  QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK TNTQENQSQS 151  NEHTGSTVGS LKGDTTIVAG KHYEQIGSTV SSPEGNNTIY AQSIDIQAAH 201  NKLNSNTTQT YEQKXLTVAF SSPVTDLAQQ ...

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the pspA Putative Secreted Protein of N. meningitidis(Accession Number AF030941)

ORF117 and pspA protein show 45% aa identity in 224aa overlap:

Orf117: 4 NLNAKAAEVSSANGTLAVSANNDINISAGINTTHVDDASKHTGRSGGGNKLVITDKAQSH63 ++  +AAEV S  G L ++A  DI + AG   T  +DA K+TGRSGGG K  +T   ++ pspA:1173 DIRIRAAEVGSEQGRLKLAAGRDIKVEAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQ 1232Orf117: 64 HETAQSSTFEGKQVVLQAGNDANILGSNVISDNGTQIQAGNHVRIGTTQTQSQSETYHQT123 +  A S T +GK+++L +G D  + GSN+I+DN T + A N++ +   +T+S+S   ++ pspA:1233 NGQAVSGTLDGKEIILVSGRDITVTGSNIIADNHTILSAKNNIVLKAAETRSRSAEMNKK 1292Orf117: 124 QKSGLM-SAGIGFTIGSKTNTQENQSQSNEHTGSTVGSLKGDTTIVAGKHYEQIGSTVSS182 +KSGLM S GIGFT GSK +TQ N+S++  HT S VGSL G+T I AGKHY Q GST+SS pspA:1293 EKSGLMGSGGIGFTAGSKKDTQTNRSETVSHTESVVGSLNGNTLISAGKHYTQTGSTISS 1352Orf117: 183 PEGNNTIYAQSIDIQAAHNKLNSNTTQTYEQKXLTVAFSSPVTD 226 P+G+  I+  I I AA N+ +  + Q YEQK +TVA S PV + pspA: 1353PQGDVGISSGKISIDAAQNRYSQESKQVYEQKGVTVAISVPVVN 1396Homology with a Predicted ORF from N. gonorrhoeae

ORF117 shows 90% identity over a 230aa overlap with a predicted ORF(ORF117ng) from N. gonorrhoeae:

An ORF117ng nucleotide sequence <SEQ ID 519> was predicted to encode aprotein having amino acid sequence <SEQ ID 520>:

1 ..LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD ETGHREQNYT 51  LPEEITRDIS LGSFAYESHS KALSRHAPSQ GTELPQSNRD NIRTAKSNGI 101  SLPYTPNSFT PLPGSSLYII NPANKGYLVE TDPRFANYRQ WLGSDYMLGS 151  LKLDPNNLHK RLGDGYYEQR LINEQIAELT GHRRLDGYQN DEEQFKALMD 201  NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLP DGGTQTVLMP 251  QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR NALIINTDTL 301  DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN INNQSTAKSS 351  QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ ISNQSDQGQT 401  RLQAGRDINL DTVQTGKYQE IHFDADNHTI RGSTNEVGSS IQTKGDVTLL 451  SGNNLNAKAA EVGSAKGTLA VYAKNDITIS SGIHAGQVDD ASKHTGRSGG 501  GNKLVITDKA QSHHETAQSS TFEGKQVVLQ AGNDANILGS NVISDNGTRI 551  QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSK TNTQENQSQS 601  NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS TQSMDIGAAQ 651  NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAK QFDKAKTTAL 701  MPWRLPMQVG RLFKQAKAPK K*

Further work revealed the following gonococcal partial DNA sequence <SEQID 521>:

1 TTGCTTGTGC AAACAGAAAA AGACGGTTTG CATAACGAGC AAACCTTTGG 51 CGAGAAGAAAGTCTTCAGCG AAAATGGTAA GTTGCACAAC TACTGGCGTG 101 CGCGTCGTAA AGGACATGATGAAACAGGGC ATCGTGAACA AAATTATACT 151 TTGCCGGAGG AAATCACACG CGACATTTCACTGGGTTCAT TTGCCTATGA 201 ATCGCATAGC AAAGCATTAA GCCGTCATGC GCCCAGCCAAGGCACTGAGT 251 TGCCACAAAG TAACCGGGAT AATATCCGTA CTGCGAAAAG CAACGGTATT301 TCGCTACCCT ATACGCCCAA TTCTTTTACC CCATTACCCG GCAGCAGCTT 351ATACATTATC AATCCTGCCA ATAAAGGCTA TCTTGTTGAA ACCGATCCAC 401 GCTTTGCCAACTACCGTCAA TGGTTGGGTA GTGACTATAT GCTGGGCAGC 451 CTCAAACTAG ACCCAAACAATTTACATAAA CGTTTGGGTG ATGGTTATTA 501 CGAGCAACGT TTAATCAATG AACAAATCGCAGAGCTGACA GGGCATCGTC 551 GTTTAGACGG TTATCAAAAC GACGAAGAAC AATTTAAAGCCTTAATGGAT 601 AATGGCGCGA CTGCGGCACG TTCGATGAAT CTCAGCGTTG GCATTGCATT651 AAGTGCCGAG CAAGCAGCGC AACTGACCAG CGATATTGTT TGGTTGGTAC 701AAAAAGAAGT TAAACTTCCT GATGGCGGCA CACAAACCGT ATTGATGCCA 751 CAGGTTTATGTACGCGTTAA AAATGGCGGC ATAGACGGTA AAGGTGCATT 801 GTTGTCAGGC AGCAATACACAAATCAATGT TTCAGGCAGC CTGAAAAACT 851 CAGGCACGAT TGCAGGGCGC AATGCGCTTATTATCAATAC CGATACGCTA 901 GACAATATCG GTGGGCGTAT TCATGCGCAA AAATCAGCGGTTACGGCCAC 951 ACAAGACATC AATAATATTG GCGGCATTCT TTCTGCCGAA CAGACATTAT1001 TGCTCAATGC GGGTAACAAC ATCAACAACC AAAGCACGGC CAAGAGCAGT 1051CAAAATGCAC AAGGTAGCAG CACCTACCTA GACCGAATGG CAGGTATTTA 1101 TATCACAGGCAAAGAAAAAG GTGTTTTAGC AGCGCAGGCA GGCAAAGACA 1151 TCAACATCAT TGCCGGTCAAATCAGCAATC AATCAGATCA AGGGCAAACC 1201 CGGCTGCAGG CAGGACGCGA CATTAACCTGGATACGGTAC AAACCGGCAA 1251 ATATCAAGAA ATCCATTTTG ATGCCGATAA CCATACCATCCGAGGTTCAA 1301 CGAACGAAGT CGGCAGCAGC ATTCAAACAA AAGGCGATGT TACCCtatTG1351 TCAGGGAATA ATCTCAATGC CAAAGCTGCC GAAGTCGGCA GCGCAAAAGG 1401CACACTTGCC GTGTATGCTA AAAATGACAT TACTATCAGC TCAGGCATCC 1451 ATGCCGGCCAAGTTGATGAT GCGTCCAAAC ATACAGGCAG AAGCGGCGGC 1501 GGTAATAAAT TAGTCATTACCGATAAAGCC CAAAGTCATC ACGAAACTGC 1551 TCAAAGCAGC ACCTTTGAAG GCAAGCAAGTTGTATTGCAG GCAGGAAACG 1601 ATGCCAACAT CCTTGGCAGT AATGTTATTT CCGATAATGGCACCCGGATT 1651 CAAGCAGGCA ATCATGTTCG CATTGGTACA ACCCAAACTC AAAGCCAAAG1701 CGAAACCTAT CATCAAACCC AAAAATCAGG ATTGATGAGT GCAGGTATCG 1751GCTTCACTAT TGGCAGCAAG ACAAACACAC AAGAAAACCA ATCCCAAAGC 1801 AACGAACATACAGGCAGTAC CGTAGGCAGC CTGAAAGGCG ATACCACCAT 1851 TGTTGCAAGC AAACACTACGAACAAACCGG CAGCAACGTT TCCAGCCCTG 1901 AGGGCAACAA CCTTATCAGC ACGCAAAGTATGGATATTGG CGCAGCACAA 1951 AACCAATTAA ACAGCAAAAC CACCCAAACC TACGAACAAAAAGGCTTAAC 2001 GGTGGCATTC AGTTCGCCCG TTACCGATTT GGCACAACAA GCGATTGCCG2051 TAGCACACAA AGCAGCAAAC AAGTCGGACA AAGCAAAAAC GACCGCGTTA 2101ATGCCATGGC GGCTGCCAAT GCAGGTTGGC AGGCCTATCA AACAGGCAAA 2151 GGCGCACAAAACTTAG

This corresponds to the amino acid sequence <SEQ ID 522; ORF117ng-1>:

1 LLVQTEKDGL HNEQTFGEKK VFSENGKLHN YWRARRKGHD ETGHREQNYT 51 LPEEITRDISLGSFAYESHS KALSRHAPSQ GTELPQSNRD NIRTAKSNGI 101 SLPYTPNSFT PLPGSSLYIINPANKGYLVE TDPRFANYRQ WLGSDYMLGS 151 LKLDPNNLHK RLGDGYYEQR LINEQIAELTGHRRLDGYQN DEEQFKALMD 201 NGATAARSMN LSVGIALSAE QAAQLTSDIV WLVQKEVKLPDGGTQTVLMP 251 QVYVRVKNGG IDGKGALLSG SNTQINVSGS LKNSGTIAGR NALIINTDTL301 DNIGGRIHAQ KSAVTATQDI NNIGGILSAE QTLLLNAGNN INNQSTAKSS 351QNAQGSSTYL DRMAGIYITG KEKGVLAAQA GKDINIIAGQ ISNQSDQGQT 401 RLQAGRDINLDTVQTGKYQE IHFDADNHTI RGSTNEVGSS IQTKGDVTLL 451 SGNNLNAKAA EVGSAKGTLAVYAKNDITIS SGIHAGQVDD ASKHTGRSGG 501 GNKLVITDKA QSHHETAQSS TFEGKQVVLQAGNDANILGS NVISDNGTRI 551 QAGNHVRIGT TQTQSQSETY HQTQKSGLMS AGIGFTIGSKTNTQENQSQS 601 NEHTGSTVGS LKGDTTIVAS KHYEQTGSNV SSPEGNNLIS TQSMDIGAAQ651 NQLNSKTTQT YEQKGLTVAF SSPVTDLAQQ AIAVAHKAAN KSDKAKTTAL 701MPWRLPMQVG RPIKQAKAHK T*

ORF117ng-1 shows the same 90% identity over a 230aa overlap with ORF117.In addition, it shows homology with a secreted N. meningitidis proteinin the database:

gi|2623258 (AF030941) putative secreted protein [Neisseria meningitidis]Length = 2273 Score = 604 bits (1541), Expect = e−172 Identities= 325/678 (47%), Positives = 449/678 (65%), Gaps = 22/678 (3%) Query: 1LLVQTEKDGLHNEQTFGEKKVFSENGKLHNYWRARRKGHDETGHREQNYTLPEEITRDIS 60 L+V T+  L N++T G K + ++ G LH Y R  +KG D TG+    Y    E++  I Sbjct: 739LIVGTPESALDNDETLGTKTI-TDKGDLHRYHRHHKKGRDSTGYSRSPYEPAPEVS-SIR 796 Query:61 LGSFAYESHSKALSRHAPSQGTELPQSNRDNIRTAKSNGISLPYTPNSFTPLPGSSLYII 120+G  AY+ +       AP Q +++P +    +     NGI   +T      LP SSL+ I Sbjct: 797MGISAYKGY-------APQQASDIPGTV---VPVVAENGIHPTFT------LPNSSLFAI 840 Query:121 NPANKGYLVETDPRFANYRQWLGSDYMLGSLKLDPNNLHKRLGDGYYEQRLINEQIAELT 180  PNKGYL+ETDP F +YR+WLGS YML +L+ DPN++HKRLGDGYYEQ+L+NEQIA+LT Sbjct: 841APNNKGYLIETDPAFTDYRKWLGSGYMLAALQQDPNHIHKRLGDGYYEQKLVNEQIAKLT 900 Query:181 GHRRLDGYQNDEEQFKALMDNGATAARSMNLSVGIALSAEQAAQLTSDIVWLVQKEVKLP 240G+RRLDGY NDEEQFKALMDNG T A+ + L+ GIALSAEQ A+LTSDIVWL  + V LP Sbjct: 901GYRRLDGYTNDEEQFKALMDNGITIAKELQLTPGIALSAEQVARLTSDIVWLENETVTLP 960 Query:241 DGGTQTVLMPQVYVRVKNGGIDGKGALLSGSNTQINVSGSLKN-SGTIAGRNALIINTDT 299 DGTQTVL P+VYVR +   ++G+GALLSGS   I  SG+++N  G IAGR ALI+N Sbjct: 961DGTTQTVLKPKVYVRARPKDMNGQGALLSGSVVDIG-SGAIENRGGLIAGREALILNAQN 1019 Query:300 LDNIGGRIHAQKSAVTATQDINNIGGILSAEQTLLLNAGNNINNQSTAKSSQNAQGSSTY 359+ N+ G +  +     A  DI N G I  AE  LLL A NNI ++S  +S+QN QGS Sbjct: 1020IKNLQGDLQGKNIFAAAGSDITNTGSI-GAENALLLKASNNIESRSETRSNQNEQGSVRN 1078 Query:360 LDRMAGIYITGKEKGVLAAQAGKDINIIAGQISNQSDQGQTRLQAGRDINLDTVQTGKYQ 419+ R+AGIY+TG++ G +   AG +I + A +++NQS+ GQT L AG DI  DT    + Q Sbjct: 1079IGRVAGIYLTGRQNGSVLLDAGNNIVLTASELTNQSEDGQTVLNAGGDIRSDTTGISRNQ 1138 Query:420 EIHFDADNHTIRGSTNEVGSSIQTKGDVTLLSGNNLNAKAAEVGSAKGTLAVYAKNDITI 479   FD+DN+ IR   NEVGS+I+T+G+++L +  ++  +AAEVGS +G L + A  DI + Sbjct: 1139NTIFDSDNYVIRKEQNEVGSTIRTRGNLSLNAKGDIRIRAAEVGSEQGRLKLAAGRDIKV 1198 Query:480 SSGIHAGQVDDASKHTGRSGGGNKLVITDKAQSHHETAQSSTFEGKQVVLQAGNDANILG 539 +G    + +DA K+TGRSGGG K  +T   ++ +  A S T +GK+++L +G D  + G Sbjct: 1199EAGKAHTETEDALKYTGRSGGGIKQKMTRHLKNQNGQAVSGTLDGKEIILVSGRDITVTG 1258 Query:540 SNVISDNGTRIQAGNHVRIGTTQTQSQSETYHQTQKSGLM-SAGIGFTIGSKTNTQENQS 598SN+I+DN T + A N++ +   +T+S+S   ++ +KSGLM S GIGFT GSK +TQ N+S Sbjct: 1259SNIIADNHTILSAKNNIVLKAAETRSRSAEMNKKEKSGLMGSGGIGFTAGSKKDTQTNRS 1318 Query:599 QSNEHTGSTVGSLKGDTTIVASKHYEQTGSNVSSPEGNNLISTQSMDIGAAQNQLNSKTT 658++  HT S VGSL G+T I A KHY QTGS +SSP+G+  IS+  + I AAQN+ + ++ Sbjct: 1319ETVSHTESVVGSLNGNTLISAGKHYTQTGSTISSPQGDVGISSGKISIDAAQNRYSQESK 1378 Query:659 QTYEQKGLTVAFSSPVTD 676 Q YEQKG+TVA S PV + Sbjct: 1379QVYEQKGVTVAISVPVVN 1396

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 63

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 523>:

1 ATGATTTACA TCGTACTGTT TCTAGCTGTC GTCCTCGCCG TTGTCGCCTA 51 CAACATGTATCAGGAAAACC AATACCGCAA AAAAGTGCGC GACCAGTTCG 101 GACACTCCGA CAAAGATGCCCTGCTCAACA GCAwAACCAG CCATGTCCGC 151 GACGGCAAAC CGTCCGGCGG GTCAGTCATGATGCCGAAAC CCCAACCGGC 201 GGTCAAAAAA ACGGCAAAAC CCCAAGACCC CGyCATGCGCAACCTGCAAG 251 AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA AGCCTCCCCG301 TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA TTATCGGCAA 351CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC GCAACGAAAC 401 CTGCCGACGCGTCGGCAAAA CCTGCACCCG TTCCGCAAAC ACCTGCAAAA 451 CCGCTGATTA CGCTCAAAGAACTGTCAAAA GTCGAATTAT CCTGGTTTGA 501 CGTGCGCATC GACTTCATCT CCTAT...

This corresponds to the amino acid sequence <SEQ ID 524; ORF119>:

1 MIYIVLFLAV VLAVVAYNMY QENQYRKKVR DQFGHSDKDA LLNSXTSHVR 51 DGKPSGGSVMMPKPQPAVKK TAKPQDPXMR NLQEQDAVYI AKQKQAKASP 101 FKTEIETALE ESGIIGNSAHTVSEPQTGHS ATKPADASAK PAPVPQTPAK 151 PLITLKELSK VELSWFDVRI DFISY...

Further work revealed the complete nucleotide sequence <SEQ ID 525>:

1 ATGATTTACA TCGTACTGTT TCTAGCTGTC GTCCTCGCCG TTGTCGCCTA 51 CAACATGTATCAGGAAAACC AATACCGCAA AAAAGTGCGC GACCAGTTCG 101 GACACTCCGA CAAAGATGCCCTGCTCAACA GCAAAACCAG CCATGTCCGC 151 GACGGCAAAC CGTCCGGCGG GTCAGTCATGATGCCGAAAC CCCAACCGGC 201 GGTCAAAAAA ACGGCAAAAC CCCAAGACCC CGCCATGCGCAACCTGCAAG 251 AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA AGCCTCCCCG301 TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA TTATCGGCAA 351CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC GCACCGAAAC 401 CTGCCGACGCGCCGGCAAAA CCTGCACCCG TTCCGCAAAC ACCTGCAAAA 451 CCGCTGATTA CGCTCAAAGAACTGTCAAAA GTCGAATTAC CCTGGTTTGA 501 CGTGCGCTTC GACTTCATCT CCTATATCGCGCTGACCGAA GCCAAAGAAC 551 TGCACGCACT GCCGCGCCTT TCCAACCGCT GCCGCTACCAGATTGTCGGC 601 TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC CGGGCATCCG651 CTATCAGGCA TTTATCGTGG GTATTCAGGC AGTCAGCCGC AACGGACTTG 701CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGTGGA CGCATTCGCA 751 CAAAGCATGGGCGGTCAGAC GCTGCACACC GACCTTGCCG CCTTTATCGA 801 AGTGGCTTCC GCACTGGACGCATTCTGCGC GCGCGTCGAC CAGACCATCG 851 CCATCCATTT GGTTTCCCCG ACCAGCATCAGCGGCGTAGA ACTGCGTTCC 901 GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCGCGTTCCACTA 951 TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG CTCAACAACG1001 AGCCGTTTAC CAACGCCCTT TTGGACAACC AGTCCTACAA AGGCTTCAGT 1051ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA CCTTCGACGA 1101 TTTGTTTATGGATTTGGCGG TACGCCTGTC CGGCCAGTTG AACCTGAATC 1151 TGGTCAACGA CAAAATGGAAGAAGTTTCGA CCCAATGGCT CAAAGACGTG 1201 CGCACTTATG TATTGGCGCG TCAGTCCGAGATGCTCAAAG TCGGTATCGA 1251 ACCGGGCGGC AAAACCGCAT TGCGCCTGTT CTCCTAA

This corresponds to the amino acid sequence <SEQ ID 526; ORF119-1>:

1 MIYIVLFLAV VLAVVAYNMY QENQYRKKVR DQFGHSDKDA LLNSKTSHVR 51 DGKPSGGSVMMPKPQPAVKK TAKPQDPAMR NLQEQDAVYI AKQKQAKASP 101 FKTEIETALE ESGIIGNSAHTVSEPQTGHS APKPADAPAK PAPVPQTPAK 151 PLITLKELSK VELPWFDVRF DFISYIALTEAKELHALPRL SNRCRYQIVG 201 CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELSAFNRQVDAFA 251 QSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP TSISGVELRS301 AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL LDNQSYKGFS 351MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME EVSTQWLKDV 401 RTYVLARQSEMLKVGIEPGG KTALRLFS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF119 shows 93.7% identity over. a 175aa overlap with an ORF (ORF119a)from strain A of N. meningitidis:

The complete length ORF119a nucleotide sequence <SEQ ID 527> is:

1 ATGATTTACA TCGTACTGTT CCTCGCCGCC GTCCTCGCCG TTGTCGCCTA 51 CAATATGTATCAGGAAAACC AATACCGCAA AAAAGTGCGC GACCAGTTCG 101 GGCACTCCGA CAAAGATGCCCTGCTCAACA GCAAAACCAG CCATGTCCGC 151 GACGGCAAAC CGTCCGGCGG GCCAGTCATGATGCCGAAAC CCCAACCGGC 201 GGTCAAAAAA ACGGCAAAAT CCCAAGACCC CGCCATGCGCAACCTGCAAG 251 AGCAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA AGCCTCCCCG301 TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAAGCGGCA TTATCGGCAA 351CTCCGCCCAC ACCGTTCCCG AACCCCAAAC CGGACATTCC GCACCAAAAC 401 CTGCCGACGCGCCGGCAAAA CCTGTTCCCG TTCCGCAAAC GCCGGCAAAA 451 CCGCTGATTA CGCTCAAAGAGCTGTCGAAG GTCGAGCTGC CCTGGTTTGA 501 CGTGCGCTTC GACTTCATCT CTTATATCGCGCTGACCGAA GCCAAAGAAC 551 TGCACGCACT GCCGCGCCTT TCCAACCGCT GCCGCTACCAGATTGTCGGC 601 TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC CGGGCATCCG651 CTATCAGGCA TTTATCGTGG GTATTCAGGC AGTCAGCCGC AACGGACTTG 701CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGTGGA TGCATTCGCA 751 CACAGCATGGGCGGTCAGAC GCTGCACACC GACCTTGCCG CCTTTATCGA 801 AGTGGCTTCC GCACTGGACGCATTCTGCGC GCGCGTCGAC CAGACTATCG 851 CCATCCATTT GGTTTCCCCG ACCAGCATCAGCGGCGTAGA ACTGCGTTCC 901 GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCGCGTTCCACTA 951 TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG CTCAACAACG1001 AGCCGTTTAC CAATGCCCTT TTGGACAACC AGTCCTATAA AGGCTTCAGT 1051ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA CCTTCGACGA 1101 TTTGTTTATGGATTTGGCGG TACGCCTGTC CGGCCAGTTG AACCTGAATC 1151 TGGTCAACGA CAAAATGGAAGAAGTTTCGA CCCAATGGCT CAAAGACGTG 1201 CGCACTTATG TATTGGCTCG TCAGTCCGAGATGCTCAAAG TCGGTATCGA 1251 ACCGGGCGGC AAAACCGCAT TGCGCCTGTT CTCCTAA

This encodes a protein having amino acid sequence <SEQ ID 528>:

1 MIYIVLFLAA VLAVVAYNMY QENQYRKKVR DQFGHSDKDA LLNSKTSHVR 51 DGKPSGGPVMMPKPQPAVKK TAKSQDPAMR NLQEQDAVYI AKQKQAKASP 101 FKTEIETALE ESGIIGNSAHTVPEPQTGHS APKPADAPAK PVPVPQTPAK 151 PLITLKELSK VELPWFDVRF DFISYIALTEAKELHALPRL SNRCRYQIVG 201 CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELSAFNRQVDAFA 251 HSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP TSISGVELRS301 AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL LDNQSYKGFS 351MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME EVSTQWLKDV 401 RTYVLARQSEMLKVGIEPGG KTALRLFS*

ORF119a and ORF119-1 show 98.6% identity in 428 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF119 shows 93.1% identity over a 175aa overlap with a predicted ORF(ORF119ng) from N. gonorrhoeae:

The complete length ORF119ng nucleotide sequence <SEQ ID 529> is:

1 ATGATTTACA TCGTACTGTT CCTCGCCGCC GTCCTCGCCG TTGTCGCCTA 51 CAATATGTATCAGGAAAACC AATACCGCAA AAAAGTGCGC GACCAGTTCG 101 GACACTCCGA CAAAGATGCCCTGCTCAACA GCAAAACCAG CCATGTCCGC 151 GACGGCAAAC CGTCCGGCGG GCCAGTCATGATGCCGAAAC CCCAACCGGC 201 GGTCAAAAAA CCGGCCAAAC CCCAAGACTC CGCCATGCGCAACCTGCAAG 251 AACAGGATGC CGTCTACATC GCCAAGCAGA AACAGGCAAA AGCCTCCCCG301 TTCAAAACCG AAATCGAAAC CGCCTTGGAA GAAATCGGCA TTATCGGCAA 351CTCCGCCCAC ACCGTTTCCG AACCCCAAAC CGGACATTCC GCACCGAAAC 401 CTGCCGACGCGCCGGCAAAA CCCGTTCCCG TTCCGCAAAC GCCGGCAAAA 451 CCGCTGATTA CGCTCAAAGAGCTGTCGAAG GTCGAGCTGC CCTGGTTTGA 501 CGTGCGCTtc gACTTCATCT CCTATATCGCGCTGACCGAA GCCAAAGAAC 551 TGCACGCACT GCCGCGCCTT tccAACCGCT GCCGCTACCAGATTGTCGGC 601 TGCACCATGG ACGACCATTT CCAGATTGCC GAACCCATCC CGGGCATCCG651 CTATCAGGCA TTTATCGTGG GTATCCAGGC AGTCAGCCGC AACGGACTTG 701CCTCGCAGGA AGAACTCTCC GCATTCAACC GCCAGGCGGA CGCATTCGCA 751 CAAAGCATGGGCGGTCAGAC GCTGCACACC GACCTTGCCG CCTTTATCGA 801 AGTGGCTTCC GCACTGGACGCATTCTGCGC GCGCGTCGAC CAGACCATCG 851 CCATCCATTT GGTTTCGCCG ACCAGCATCAGCGGCGTAGA ACTGCGTTCC 901 GCCGTAACGG GCGTGGGTTT CGTTTTGGAA GACGACGGCGCGTTCCACTA 951 TACCGACACG TCGGGCTCGA CCATGTTCTC CATCTGCTCG CTCAACAACG1001 AGCCGTTTAC CAATGCCCTT TTGGACAACC AGTCCTACAA AGGCTTCAGT 1051ATGCTGCTCG ACATCCCGCA CTCTCCGGCA GGCGAAAAAA CCTTCGACGA 1101 TTTGTTTATGGATTTGGCGG TACGCCTGTC CGGTCAGTTG AACCTGAATC 1151 TGGTCAACGA CAAAATGGAAGAAGTTTCGA CCCAATGGCT CAAAGACGTA 1201 CGCACTTATG TATTGGCGCG TCAGTCCGAGATGCTCAAAG TCGGTATCGA 1251 ACCGGGCGGC AAAACCGCCC TGCGCCTGTT TTCATAA

This encodes a protein having amino acid sequence <SEQ ID 530>:

1 MIYIVLFLAA VLAVVAYNMY QENQYRKKVR DQFGHSDKDA LLNSKTSHVR 51 DGKPSGGPVMMPKPQPAVKK PAKPQDSAMR NLQEQDAVYI AKQKQAKASP 101 FKTEIETALE EIGIIGNSAHTVSEPQTGHS APKPADAPAK PVPVPQTPAK 151 PLITLKELSK VELPWFDVRF DFISYIALTEAKELHALPRL SNRCRYQIVG 201 CTMDDHFQIA EPIPGIRYQA FIVGIQAVSR NGLASQEELSAFNRQADAFA 251 QSMGGQTLHT DLAAFIEVAS ALDAFCARVD QTIAIHLVSP TSISGVELRS301 AVTGVGFVLE DDGAFHYTDT SGSTMFSICS LNNEPFTNAL LDNQSYKGFS 351MLLDIPHSPA GEKTFDDLFM DLAVRLSGQL NLNLVNDKME EVSTQWLKDV 401 RTYVLARQSEMLKVGIEPGG KTALRLFS*

ORF119ng and ORF119-1 show 98.4% identity over 428 aa overlap:

Based on this analysis, including the presence of a putative leadersequence in the gonococcal protein, it is predicted that the proteinsfrom N. meningitidis and N. gonorrhoeae, and their epitopes, could beuseful antigens for vaccines or diagnostics, or for raising antibodies.

Example 64

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 531>

1 ..GCGCGGCACG GCACGGAAGA TTTCTTCATG AACAACAGCG ACAC.ATCAG 51  GCAGATAGTC GAAAGCACCA CCGGTACGAT GAAGCTGCTG ATTTCCTCCA 101  TCGCCCTGAT TTCATTGGTA GTCGGCGGCA TCGGCGTGAT GAACATCATG 151  CTGGTGTCCG TTACCGAGCG CACCAAAGAA ATCGGCATAC GGATGGCAAT 201  CGGCGCGCGG CGCGGCAATA TTTyGCAGCA GTTTTTGATT GAGGCGGTGT 251  TAATCTGCGT CATCGGCGGT TTGGTCGGCG TGGGTTTGTC CGCCGCCGTC 301  AGCCTCGTGT TCAATCATTT TGTAACCGAC TTCCCGATGG ACATTTCCGC 351  CATGTCCGTC ATCGGCGCGG TCGCCTGTTC GACCGGAATC GGCATCGCGT 401  TCGGCTTTAT GCCTGCCAAT AAAGCAGCCA AACTCAATCC GATAGACGCA 451  TTGGCACAGG ATTGA

This corresponds to the amino acid sequence <SEQ ID 532; ORF134>:

1 ..ARHGTEDFFM NNSDXIRQIV ESTTGTMKLL ISSIALISLV VGGIGVMNIM 51  LVSVTERTKE IGIRMAIGAR RGNIXQQFLI EAVLICVIGG LVGVGLSAAV 101  SLVFNHFVTD FPMDISAMSV IGAVACSTGI GIAFGFMPAN KAAKLNPIDA 151   LAQD*

Further work revealed the complete nucleotide sequence <SEQ ID 533>:

1 ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC TTCTGACGAT 51 GCTCGGCATCATCATCGGTA TCGCGTCGGT GGTTTCCGTC GTCGCATTGG 101 GCAATGGTTC GCAGAAAAAAATCCTTGAAG ACATCAGTTC GATAGGGACG 151 AACACCATCA GCATCTTCCC GGGGCGCGGCTTCGGCGACA GGCGCAGCGG 201 CAGGATTAAA ACCCTGACCA TAGACGACGC AAAAATCATCGCCAAACAAA 251 GCTACGTTGC TTCCGCCACG CCCATGACTT CGAGCGGCGG CACGCTGACT301 TACCGCAACA CCGACCTGAC CGCCTCGCTT TACGGCGTGG GCGAACAATA 351TTTCGACGTG CGCGGACTGA AGCTGGAAAC GGGGCGGCTG TTTGACGAAA 401 ACGATGTGAAAGAAGACGCG CAGGTCGTCG TCATCGACCA AAATGTCAAA 451 GACAAACTCT TTGCGGACTCGGATCCGTTG GGTAAAACCA TTTTGTTCAG 501 GAAACGCCCC TTGACCGTCA TCGGCGTGATGAAAAAAGAC GAAAACGCTT 551 TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATACGACGGTGATG 601 CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG TCAAAATCAA651 AGACAATGCC AATACCCAGG TTGCCGAAAA AGGGCTGACC GATCTGCTCA 701AAGCGCGGCA CGGCACGGAA GATTTCTTCA TGAACAACAG CGACAGCATC 751 AGGCAGATAGTCGAAAGCAC CACCGGTACG ATGAAGCTGC TGATTTCCTC 801 CATCGCCCTG ATTTCATTGGTAGTCGGCGG CATCGGCGTG ATGAACATCA 851 TGCTGGTGTC CGTTACCGAG CGCACCAAAGAAATCGGCAT ACGGATGGCA 901 ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGATTGAGGCGGT 951 GTTAATCTGC GTCATCGGCG GTTTGGTCGG CGTGGGTTTG TCCGCCGCCG1001 TCAGCCTCGT GTTCAATCAT TTTGTAACCG ACTTCCCGAT GGACATTTCC 1051GCCATGTCCG TCATCGGCGC GGTCGCCTGT TCGACCGGAA TCGGCATCGC 1101 GTTCGGCTTTATGCCTGCCA ATAAAGCAGC CAAACTCAAT CCGATAGACG 1151 CATTGGCACA GGATTGA

This corresponds to the amino acid sequence <SEQ ID 534; ORF134-1>:

1 MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK ILEDISSIGT 51 NTISIFPGRGFGDRRSGRIK TLTIDDAKII AKQSYVASAT PMTSSGGTLT 101 YRNTDLTASL YGVGEQYFDVRGLKLETGRL FDENDVKEDA QVVVIDQNVK 151 DKLFADSDPL GKTILFRKRP LTVIGVMKKDENAFGNSDVL MLWSPYTTVM 201 HQITGESHTN SITVKIKDNA NTQVAEKGLT DLLKARHGTEDFFMNNSDSI 251 RQIVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE RTKEIGIRMA301 IGARRGNILQ QFLIEAVLIC VIGGLVGVGL SAAVSLVFNH FVTDFPMDIS 351AMSVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Hypothetical Protein o648 of E. coli (Accession NumberAE000189)

ORF134 and o648 protein show 45% aa identity in 153aa overlap:

Orf134:   2 RHGTEDFFMNNSDXIRQIVESTTGTMKXXXXXXXXXXXVVGGIGVMNIMLVSVTERTKEI  61            RHG +DFFN   D + + VE TT T++           VVGGIGVMNIMLVSVTERT+EIo648:   496 RHGKKDFFTWNMDGVLKTVEKTTRTLQLFLTLVAVISLVVGGIGVMNIMLVSVTERTREI 555Orf134:  62 GIRMAIGARRGNIXQQFLIEAXXXXXXXXXXXXXXXXXXXXXFNHFVTDFPMDISAMSVI 121            GIRMA+GAR  ++ QQFLIEA                        F+ + +   S ++++o648:   556 GIRMAVGARASDVLQQFLIEAVLVCLVGGALGITLSLLIAFTLQLFLPGWEIGFSPLALL 615Orf134: 122 GAVACSTGIGIAFGFMPANKAAKLNPIDALAQD                            154             A  CST  GI FG++PA  AA+L+P+DALA++o648:   616 LAFLCSTVTGILFGWLPARNAARLDPVDALARE                            648Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF134 shows 98.7% identity over a 154aa overlap with an ORF (ORF134a)from strain A of N. meningitidis:

The complete length ORF134a nucleotide sequence <SEQ ID 535> is:

1 ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC TTCTGACGAT 51GCTCGGCATC ATCATCGGTA TCGCTTCGGT TGTCTCCGTC GTCGCATTGG 101GCAACGGTTC GCAGAAAAAA ATCCTTGAAG ACATCAGTTC GATAGGGACG 151AACACCATCA GCATCTTCCC AGGGCGCGGC TTCGGCGACA GGCGCAGCGG 201CAGGATTAAA ACCCTGACCA TAGACGACGC AAAAATCATC GCCAAACAAA 251GCTACGTTGC TTCCGCCACG CCCATGACTT CGAGCGGCGG CACGCTGACT 301TACCGCAATA CCGACCTGAC CGCTTCTTTG TACGGTGTGG GCGAACAATA 351TTTCGACGTG CGCGGGCTGA AGCTGGAAAC GGGGCGGCTG TTTGACGAAA 401ACGATGTGAA AGAAGACGCG CAGGTCGTCG TCATCGACCA AAATGTCAAA 451GACAAACTCT TTGCGGACTC GGATCCGTTG GGTAAAACCA TTTTGTTCAG 501GAAACGCCCC TTGACCGTCA TCGGCGTGAT GAAAAAAGAC GAAAACGCTT 551TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATAC GACGGTGATG 601CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG TCAAAATCAA 651AGACAATGCC AATACCCAGG TTGCCGAAAA AGGGCTGACC GATCTGCTCA 701AAGCGCGGCA CGGCACGGAA GATTTCTTCA TGAACAACAG CGACAGCATC 751AGGCAGATAG TCGAAAGCAC CACCGGTACG ATGAAGCTGC TGATTTCCTC 801CATCGCCCTG ATTTCATTGG TAGTCGGCGG CATCGGCGTG ATGAACATCA 851TGCTGGTGTC CGTTACCGAG CGCACCAAAG AAATCGGCAT ACGGATGGCA 901ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGA TTGAGGCGGT 951GTTAATCTGC GTCATCGGCG GTTTGGTCGG CGTGGGTTTG TCCGCCGCCG 1001TCAGCCTCGT GTTCAATCAT TTTGTAACCG ACTTCCCGAT GGACATTTCC 1051GCCATGTCCG TCATCGGCGC GGTCGCCTGT TCGACCGGAA TCGGCATCGC 1101GTTCGGCTTT ATGCCTGCCA ATAAAGCAGC CAAACTCAAT CCGATAGATG 1151CATTGGCGCA GGATTGA

This encodes a protein having amino acid sequence <SEQ ID 536>:

1 MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK ILEDISSIGT 51NTISIFPGRG FGDRRSGRIK TLTIDDAKII AKQSYVASAT PMTSSGGTLT 101YRNTDLTASL YGVGEQYFDV RGLKLETGRL FDENDVKEDA QVVVIDQNVK 151DKLFADSDPL GKTILFRKRP LTVIGVMKKD ENAFGNSDVL MLWSPYTTVM 201HQITGESHTN SITVKIKDNA NTQVAEKGLT DLLKARHGTE DFFMNNSDSI 251RQIVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE RTKEIGIRMA 301IGARRGNILQ QFLIEAVLIC VIGGLVGVGL SAAVSLVFNH FVTDFPMDIS 351AMSVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

ORF134a and ORF134-1 show 100.0% identity in 388 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF134 shows 96.8% identity over a 154aa overlap with a predicted ORF(ORF 134.ng) from N. gonorrhoeae:

The complete length ORF134ng nucleotide sequence <SEQ ID 537> is:

1 ATGTCGGTGC AAGCAGTATT GGCGCACAAA ATGCGTTCGC TTCTGACCAT 51GCTCGGCATC ATCATCGGTA TCGCTTCGGT TGTCTCCGTC GTCGCGCTGG 101GCAACGGTTC GCAGAAAAAA ATCCTCGAAG ACATCAGTTC GATGGGGACG 151AACACCATCA GCATCTTCCC CGGGCGCGGC TTCGGCGACA GGCGCAGCGG 201CAAAATCAAA ACCCTGACCA TAGACGACGC AAAAATCATC GCCAAACAAA 251GCTACGTTGC CTCCGCCACG CCCATGACTT CGAGCGGCGG CACGCTGACC 301TACCGCAATA CCGACCTGAC CGCTTCTTTG TACGGTGTGG GCGAACAATA 351TTTCGACGTG CGCGGGCTGA AGCTGGAAAC GGGGCGGCTG TTTGATGAGA 401ACGATGTGAA AGAAGACGCG CAAGTCGTCG TCATCGACCA AAATGTCAAA 451GACAAACTCT TTGCGGACTC GGATCCGTTG GGTAAAACCA TTTTGTTCAG 501GAAACGCCCC TTGACCGTCA TCGGCGTGAT GAAAAAAGAC GAAAACGCTT 551TCGGCAATTC CGACGTGCTG ATGCTTTGGT CGCCCTATAC GACGGTGATG 601CACCAAATCA CAGGCGAGAG CCACACCAAC TCCATCACCG TCAAAATCAA 651AGACAATGCC AATACCCGGG TTGCCGAAAA AGGGCTGGCC GAGCTGCTCA 701AAGCACGGCA CGGCACGGAA GACTTCTTTA TGAACAACAG CGACAGCATC 751AGGCAGATGG TCGAAAGCAC CACCGGTACG ATGAAGCTGC TGATTTCCTC 801CATCGCCCTG ATTTCATTGG TAGTCGGCGG CATCGGTGTG ATGAACATTA 851TGCTGGTGTC CGTTACCGAG CGCACCAAAG AAATCGGCAT ACGGATGGCA 901ATCGGCGCGC GGCGCGGCAA TATTTTGCAG CAGTTTTTGA TTGAGGCGGT 951GTTAATCTGC ATCATCGGAG GCTTGGTCGG CGTAGGTTTG TCCGCCGCCG 1001TCAGCCTCGT GTTCAATCAT TTTGTAACCG ATTTCCCGAT GGACATTTCG 1051GCGGCATCCG TTATCGGGGC GGTCGCCTGT TCGACCGGAA TCGGCATCGC 1101GTTCGGCTTT ATGCCTGCCA ATAAGGCAGC CAAACTCAAT CCGATAGATG 1151CATTGGCGCA GGATTGA

This encodes a protein having amino acid sequence <SEQ ID 538>:

1 MSVQAVLAHK MRSLLTMLGI IIGIASVVSV VALGNGSQKK ILEDISSMGT 51NTISIFPGRG FGDRRSGKIK TLTIDDAKII AKQSYVASAT PMTSSGGTLT 101YRNTDLTASL YGVGEQYFDV RGLKLETGRL FDENDVKEDA QVVVIDQNVK 151DKLFADSDPL GKTILFRKRP LTVIGVMKKD ENAFGNSDVL MLWSPYTTVM 201HQITGESHTN SITVKIKDNA NTRVAEKGLA ELLKARHGTE DFFMNNSDSI 251RQMVESTTGT MKLLISSIAL ISLVVGGIGV MNIMLVSVTE RTKEIGIRMA 301IGARRGNILQ QFLIEAVLIC IIGGLVGVGL SAAVSLVFNH FVTDFPMDIS 351AASVIGAVAC STGIGIAFGF MPANKAAKLN PIDALAQD*

ORF134ng and ORF134-1 show 97.9% identity in 388 aa overlap:

ORF134ng also shows homology to an E. coli ABC transporter:

sp|P75831|YBJZ_ECOLI HYPOTHETICAL ABC TRANSPORTER ATP-BINDINGPROTEIN YBJZ >gi5 (AE000189) o648; similar to YBBA_HAEIN SW: P45247[Escherichia coli] Length = 648 Score = 297 bits (753), Expect = 6e−80Identities = 162/389 (41%), Positives = 230/389 (58%), Gaps = 1/389 (0%)Query: 1 MSVQAVLAHKMRSLLTMLXXXXXXXXXXXXXXLGNGSQKKILEDISSMGTNTISIFPGRG 60M+ +A+ A+KMR+LLTML              +G+ +++ +L DI S+GTNTI ++PG+ Sbjct: 260MAWRALAANKMRTLLTMLGIIIGIASVVSIVVVGDAAKQMVLADIRSIGTNTIDVYPGKD 319 Query:61 FGDRRSGKIKTLTIDDAKIIAKQSYVASATPMTSSGGTLTYRNTDLTASLYGVGEQYFDV 120FGD      + L  DD   I KQ +VASATP  S    L Y N D+ AS  GV   YF+V Sbjct: 320FGDDDPQYQQALKYDDLIAIQKQPWVASATPAVSQNLRLRYNNVDVAASANGVSGDYFNV 379 Query:121 RGLKLETGRLFDENDVKEDAQVVVIDQNVKDKLFAD-SDPLGKTILFRKRPLTVIGVMKK 179  G+   G  F++  +   AQVVV+D N + +LF   +D +G+ IL    P  VIGV ++ Sbjct: 380YGMTFSEGNTFNQEQLNGRAQVVVLDSNTRRQLFPHKADVVGEVILVGNMPARVIGVAEE 439 Query:180 DENAFGNSDVLMLWSPYTTVMHQITGESHTNSITVKIKDNANTRVAEKGLAELLKARHGT 239  ++FG+S VL +W PY+T+  ++ G+S  NSITV++K+  ++  AE+ L  LL  REG Sbjct: 440KQSMFGSSKVLRVWLPYSTMSGRVMGQSWLNSITVRVKEGFDSAEAEQQLTRLLSLRHGK 499 Query:240 EDFFMNNSDSIRQMVESTTGTMKXXXXXXXXXXXVVGGIGVMNIMLVSVTERTKEIGIRM 299+DFF  N D + + VE TT T++           VVGGIGVMNIMLVSVTERT+EIGIRM Sbjct: 500KDFFTWNMDGVLKTVEKTTRTLQLFLTLVAVISLVVGGIGVMNIMLVSVTERTREIGIRM 559 Query:300 AIGARRGNILQQFLIEXXXXXXXXXXXXXXXXXXXXXXFNHFVTDFPMDISAASVIGAVA 359A+GAR  ++LQQFLIE                         F+  + +  S  +++ A Sbjct: 560AVGARASDVLQQFLIEAVLVCLVGGALGITLSLLIAFTLQLFLPGWEIGFSPLALLLAFL 619 Query:360 CSTGIGIAFGFMPANKAAKLNPIDALAQD 388 CST  GI FG++PA  AA+L+P+DALA++Sbjct: 620 CSTVTGILFGWLPARNAARLDPVDALARE 648

Based on this analysis, including the presence of the leader peptide andtransmembrane regions in the gonococcal protein, it is prediceted thatthese proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 65

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 539>:

1 ..GGGACGGGAG CGATGCTGCT GCTGTTTTAC GCGGTAACGA T.CTGCCTTT 51  GGCCACTGGC GTTACCCTGA GTTACACCTC GTCGATTTTT TTGGCGGTAT 101  TTTCCTTCCT GATTTTGAAA GAACGGATTT CCGTTTACAC GCAGGCGGTG 151  CTGCTCCTTG GTTTTGCCGG CGTGGTATTG CTGCTTAATC CCTCGTTCCG 201  CAGCGGTCAG GAAACGGCGG CACTCGCCGG GCTGGCGGGC GGCGCGATGT 251  CCGGCTGGGC GTATTTGAAA GTGCGCGAAC TGTCTTTGGC GGGCGAACCC 301  GGCTGGCGCG TCGTGTTTTA CCTTTCCGTG ACAGGTGTGG CGATGTCGTC 351  GGTTTGGGCG ACGCTGACCG GCTGGCACAC CCTGTCCTTT CCATCGGCAG 401  TTTATCTGTC GTGCATCGGC GTGTCCGCGC TGATTGCCCA ACTGTCGATG 451  ACGCGCGCCT ACAAAGTCGG CGACAAATTC ACGGTTGCCT CGCTTTCCTA 501  TATGACCGTC GTTTTTTCCG CTCTGTCTGC CGCATTTTTT CTGGGCGAAG 551  AGCTTTTCTG GCAGGAAATA CTCGGTATGT GCATCATCAT CCTCAGCGGT 601   ATTTTGA

This corresponds to the amino acid sequence <SEQ ID 540; ORF135>:

1 ..GTGAMLLLFY AVTILPLATG VTLSYTSSIF LAVFSFLILK ERISVYTQAV 51  LLLGFAGVVL LLNPSFRSGQ ETAALAGLAG GAMSGWAYLK VRELSLAGEP 101  GWRVVFYLSV TGVAMSSVWA TLTGWHTLSF PSAVYLSCIG VSALIAQLSM 151  TRAYKVGDKF TVASLSYMTV VFSALSAAFF LGEELFWQEI LGMCIIISAV 201   F*

Further work revealed the complete nucleotide sequence <SEQ ID 541>:

1 ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA TGCTGGTGGC 51GGCGGCCTGC TTTACCATTA TGAACGTATT GATTAAAGAG GCATCGGCAA 101AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT GCTGTTTTCA 151ACCGTTGCGC TCGGGGCTGC CGCCGTATTG CGTCGGGACA mCTTCCGCAC 201GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG ACGGGGGCGA 251TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGGC CACTGGCGTT 301ACCCTGAGTT ACACCTCGTC GATTTTTTTG GCGGTATTTT CCTTCCTGAT 351TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG CTCCTTGGTT 401TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG CGGTCAGGAA 451ACGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG GCTGGGCGTA 501TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC TGGCGCGTCG 551TGTTTTACCT TTCCGTGACA GGTGTGGCGA TGTCGTCGGT TTGGGCGACG 601CTGACCGGCT GGCACACCCT GTCCTTTCCA TCGGCAGTTT ATCTGTCGTG 651CATCGGCGTG TCCGCGCTGA TTGCCCAACT GTCGATGACG CGCGCCTACA 701AAGTCGGCGA CAAATTCACG GTTGCCTCGC TTTCCTATAT GACCGTCGTT 751TTTTCCGCTC TGTCTGCCGC ATTTTTTCTG GGCGAAGAGC TTTTCTGGCA 801GGAAATACTC GGTATGTGCA TCATCATCCT CAGCGGTATT TTGAGCAGCA 851TCCGCCCCAC TGCCTTCAAA CAGCGGCTGC AATCCCTGTT CCGCCAAAGA 901 TAA

This corresponds to the amino acid sequence <SEQ ID 542; ORF135-1>:

1 MDTAKKDILG SGWMLVAAAC FTIMNVLIKE ASAKFALGSG ELVFWRMLFS 51TVALGAAAVL RRDXFRTPHW KNHLNRSMVG TGAMLLLFYA VTHLPLATGV 101TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL LNPSFRSGQE 151TAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSVT GVAMSSVWAT 201LTGWHTLSFP SAVYLSCIGV SALIAQLSMT RAYKVGDKFT VASLSYMTVV 251FSALSAAFFL GEELFWQEIL GMCIIILSGI LSSIRPTAFK QRLQSLFRQR 301 *

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF135 shows 99.0% identity over a 197aa overlap with an ORF (ORF135a)from strain A of N. meningitidis:

The complete length ORF135a nucleotide sequence <SEQ ID 543> is:

1 ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA TGCTGGTGGC 51GGCGGCCTGC TTTACCATTA TGAACGTATT GATTAAAGAG GCATCGGCAA 101AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT GCTGTTTTCA 151ACCGTTGCGC TCGGGGCTGC CGCCGTATTG CGTCGGGACA CCTTCCGCAC 201GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG ACGGGGGCGA 251TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGGC CACCGGCGTT 301ACCCTGAGTT ACACCTCGTC GATTTTTTTG GCGGTATTTT CCTTCCTGAT 351TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG CTCCTTGGTT 401TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG CGGTCAGGAA 451ACGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG GCTGGGCGTA 501TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC TGGCGCGTCG 551TGTTTTACCT TTCCGTGACA GGTGTGGCGA TGTCATCGGT TTGGGCGACG 601CTGACCGGCT GGCACACCCT GTCCTTTCCA TCGGCAGTTT ATCTGTCGTG 651CATCGGCGTG TCCGCGCTGA TTGCCCAACT GTCGATGACG CGCGCCTACA 701AAGTCGGCGA CAAATTCACG GTTGCCTCGC TTTCCTATAT GACCGTCGTT 751TTTTCCGCTC TGTCTGCCGC ATTTTTTCTG GCCGAAGAGC TTTTCTGGCA 801GGAAATACTC GGTATGTGCA TCATCATCCT CAGCGGTATT TTGAGCAGCA 851TCCGCCCCAC TGCCTTCAAA CAGCGGCTGC AATCCCTGTT CCGCCAAAGA 901 TAA

This encodes a protein having amino acid sequence <SEQ ID 544>:

1 MDTAKKDILG SGWMLVAAAC FTIMNVLIKE ASAKFALGSG ELVFWRMLFS 51TVALGAAAVL RRDTFRTPHW KNHLNRSMVG TGAMLLLFYA VTHLPLATGV 101TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL LNPSFRSGQE 151TAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSVT GVAMSSVWAT 201LTGWHTLSFP SAVYLSCIGV SALIAQLSMT RAYKVGDKFT VASLSYMTVV 251FSALSAAFFL AEELFWQEIL GMCIIILSGI LSSIRPTAFK QRLQSLFRQR 301 *

ORF135a and ORF135-1 show 99.3% identity in 300 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF135 shows 97% identity over a 201aa overlap with a predicted ORF(ORF135ng) from N. gonorrhoeae:

An ORF135ng nucleotide sequence <SEQ ID 545> was predicted to encode aprotein having amino acid sequence <SEQ ID 546>:

1 MPSEKAFRRH LRTASFQGLH LHHFHQKVGK CGIIGFGIHI FPTLLPAAQG 51ILDIQLGLFR IDFAALAVYR RTQVDFIHTV IDGIASDQAF SEVVQILRRL 101NLGHFTDTHL IAQARRFIAD FGNIRPMRRG EAKTFCRCFR FDGIDGIHGD 151FRQCGHINRL APGKDCRNGK RDKVFFHTRH YNQVCLEKTN CSARKIKFRH 201QKQAKTHSTS LAARFTIRPS LSQRPFMDTA KKDILGSGWM LVAAACFTVM 251NVLIKEASAK FALGSGELVF WRMLFSTVTL GAAAVLRRDT FRTPHWKNHL 301NRSMVGTGAM LLLFYAVTHL PLTTGVTLSY TSSIFLAVFS FLILKERISV 351YTQAVLLLGF AGVVLLLNPS FRSGQEPAAL AGLAGGAMSG WAYLKVRELS 401LAGEPGWRVV FYLSATGVAM SSVWATLTGW HTLSFPSAVY LSGIGVSALI 451AQLSMTRAYK VGDKFTVASL SYMTVVFSAL SAAFFLGEEL FWQEILGMCI 501 IISAAF*

Further work revealed the following gonococcal sequence <SEQ ID 547>:

1 ATGGATACCG CAAAAAAAGA CATTTTAGGA TCGGGCTGGA TGCTGGTGGC 51GGCGGCCTGC TTCACCGTTA TGAACGTATT GATTAAAGAG GCATCGGCAA 101AATTTGCCCT CGGCAGCGGC GAATTGGTCT TTTGGCGCAT GCTGTTTTCA 151ACCGTTACGC TCGGTGCTGC CGCCGTATTG CGGCGCGACA CCTTCCGCAC 201GCCCCATTGG AAAAACCACT TAAACCGCAG TATGGTCGGG ACGGGGGCGA 251TGCTGCTGCT GTTTTACGCG GTAACGCATC TGCCTTTGAC AACCGGCGTT 301ACCCTGAGTT ACACCTCGTC GATTTTTttg GCGGTATTTT CCTTCCTCAT 351TTTGAAAGAA CGGATTTCCG TTTACACGCA GGCGGTGCTG CTCCTTGGTT 401TTGCCGGCGT GGTATTGCTG CTTAATCCCT CGTTCCGCAG CGGTCAGGAA 451CCGGCGGCAC TCGCCGGGCT GGCGGGCGGC GCGATGTCCG GCTGGGCGTA 501TTTGAAAGTG CGCGAACTGT CTTTGGCGGG CGAACCCGGC TGGCGCGTCG 551TGTTTTACCT TTCCGCAACC GGCGTGGCGA TGTCGTCggt ttgggcgacg 601Ctgaccggct ggCACAcccT GTCCTTTcca tcggcagttt ATCtgtCGGG 651CATCGGCGTG tccgcgCtgA TTGCCCAaCT GtcgatgAcg cGCGcctaca 701aaGTCGGCGA CAAATTCACG GTTGCCTCGC tttcctaTAt gaccgtcGTC 751TTTTCCGCCC TGTCTGCCGC ATTTTTTCTg ggcgaagagc tttTCtggCA 801GGAAATACTC GGTATGTGCA TCATTAtccT CAGCGGCATT TTGAGCAGCA 851TCCGCCCCAT TGCCTTCAAA CAGCGGCTGC AAGCCCTCTT CCGCCAAAGA 901 TAA

This corresponds to the amino acid sequence <SEQ ID 548; ORF135ng-1>:

1 MDTAKKDILG SGWMLVAAAC FTVMNVLIKE ASAKFALGSG ELVFWRMLFS 51TVTLGAAAVL RRDTFRTPHW KNHLNRSMVG TGAMLLLFYA VTHLPLTTGV 101TLSYTSSIFL AVFSFLILKE RISVYTQAVL LLGFAGVVLL LNPSFRSGQE 151PAALAGLAGG AMSGWAYLKV RELSLAGEPG WRVVFYLSAT GVAMSSVWAT 201LTGWHTLSFP SAVYLSGIGV SALIAQLSMT RAYKVGDKFT VASLSYMTVV 251FSALSAAFFL GEELFWQEIL GMCIIILSGI LSSIRPIAFK QRLQALFRQR 301 *

ORF135ng-1 and ORF135-1 show 97.0% identity in 300 aa overlap:

Based on this analysis, including the presence of several putativetransmembrane domains in the gonococcal protein, it is predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 66

The following DNA sequence was identified in N. meningitidis <SEQ ID549>:

1 ATGAAGCGGC GTATAGCCGT CTTCGTCCTG TTCCCGCAGA TAATCCGAGT 51TTTGGGACAA CTGTTGCCGA AAATCGTCAA TACAGTTCCG GCACATCGGA 101TGCTCTTCCA GATTTTCGGG ATGTTCTTTT TCTTCATACA CCAGCAATAT 151CTGCCCGGGA TCGCCGAAAT CGATTCCCCA TGCGGCATCG TGTTCGGTGC 201GCTCCTCTTC CGTCATCTGC CCGCGCATTG CCTGTATGGT AAAGCCGCCG 251TAGGGGATGC CgTTGCACAC GAACATCCAG TCGCTGATGT CGTCAACCGG 301AACGCAAACG cTTTCGCCTT GTTCGACATT GGTCAGTTCG CCsGGTTCAT 351TGTTCAGCAC ACCGTAAATA TAAAGACCGT CAAAATAAAT ATCGTCGATC 401CACATATGTT CGCAAATTTC GCCGTCTTCG CCGTCTTGGA AAAAAGGGAC 451TTTGACCATG GCAAAATCCA AGGCGGAAAT AATGCGGCGG CGTTCCCAAA 501AAAGcTCGCG CCAAAAATAT TTGAATGTTT TACGGGCGCG TTCGTCGGCA 551CGGTTTACCG GTTCGTCTGC CTGTTCTACA TAATAAATGA CGGAATCGCC 601CATCATATCT GCTCCTCAAC GTGTACGGTA TCTGTTTGCA CCTTACTGCG 651GCTTTCTgcC kTCGGCATCC GATTCGGATT TGAAAAGTTC mmrwyATTCG 701 GAATAG

This corresponds to the amino acid sequence <SEQ ID 550; ORF136>:

1 MKRRIAVFVL FPQIIRVLGQ LLPKIVNTVP AHRMLFQIFG MFFFFIHQQY 51 LPGIAEIDSPCGIVFGALLF RHLPAHCLYG KAAVGDAVAH EHPVADVVNR 101 NANAFALFDI GQFAXFIVQHTVNIKTVKIN IVDPHMFANF AVFAVLEKRD 151 FDHGKIQGGN NAAAFPKKLA PKIFECFTGAFVGTVYRFVC LFYIINDGIA 201 HHSAPQRVRY LFAPYCGFLP SASDSDLKSS XXSE*

Further work revealed the complete nucleotide sequence <SEQ ID 551>:

1 ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGTTCCCGC AGATAATCCG 51 AGTTTTGGGACAACTGTTGC CGAAAATCGT CAATACAGTT CCGGCACATC 101 GGATGCTCTT CCAGATTTTCGGGATGTTCT TTTTCTTCAT ACACCAGCAA 151 TATCTGCCCG GGATCGCCGA AATCGATTCCCCATGCGGCA TCGTGTTCGG 201 TGCGCTCCTC TTCCGTCATC TGCCCGCGCA TTGCCTGTATGGTAAAGCCG 251 CCGTAGGGGA TGCCGTTGCA CACGAACATC CAGTCGCTGA TGTCGTCAAC301 CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT TCGCCGGGTT 351CATTGTTCAG CACACCGTAA ATATAAAGAC CGTCAAAATA AATATCGTCG 401 ATCCACATATGTTCGCAAAT TTCGCCGTCT TCGCCGTCTT GGAAAAAAGG 451 GACTTTGACC ATGGCAAAATCCAAGGCGGA AATAATGCGG CGGCGTTCCC 501 AAAAAAGCTC GCGCCAAAAA TATTTGAATGTTTTACGGGC GCGTTCGTCG 551 GCACGGTTTA CCGGTTCGTC TGCCTGTTCT ACATAATAAATGACGGAATC 601 GCCCATCATT CTGCTCCTCA ACGTGTACGG TATCTGTTTG CACCTTACTG651 CGGCTTTCTG CCTTCGGCAT CCGATTCGGA TTTGAAAAGT TCCAAATATT 701 CGGAATAG

This corresponds to the amino acid sequence <SEQ ID 552; ORF136-1>:

1 MMKRRIAVFV LFPQIIRVLG QLLPKIVNTV PAHRMLFQIF GMFFFFIHQQ 51 YLPGIAEIDSPCGIVFGALL FRHLPAHCLY GKAAVGDAVA HEHPVADVVN 101 RNANAFALFD IGQFAGFIVQHTVNIKTVKI NIVDPHMFAN FAVFAVLEKR 151 DFDHGKIQGG NNAAAFPKKLAPKIFECFTG AFVGTVYRFV CLFYIINDGI 201 AHHSAPQRVR YLFAPYCGFL PSASDSDLKSSKYSE*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF136 shows 71.7% identity over. a 237aa overlap with an ORF (ORF136a)from strain A of N. meningitidis:

The complete length ORF136a nucleotide sequence <SEQ ID 553> is:

1 ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGCTCATGC AGAAAATCCG 51 GATTTTGGGACAACTGTTGC CGAAAATCGT CAATACAGTT CCGGCACATC 101 GGATGCTCTT CCAGATNTTCGGGATGTTCT TTTTCTTCAT ACACCAGCAA 151 TACCTGCCCG GGATCGCCGA AATCGATTCCCCATGCGGCA TCGTGTTCGG 201 TACGCTCCTC TTCCGTCATC NGTCCACGCA TTGCCTGTATGGTAAAGCCG 251 CCGTAGGGAA TGCCGTTGCA CACGAACATC CAGTCGCTGA TGTCGTCAAC301 CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT TCGCCGGGTT 351CATTGTTCAG CACGCCATAA ATGTAAAGAC CGTCAAAATA AATATCGTCG 401 ATCCACATATGTTCGCAAAT TTCGCCNTCT TCGCCGTCTT GGAAAAAAGG 451 GCTTTGACCA TGGCAAAATCTAAGGNGNNA NNGATGCGGC GGCGTTCCCA 501 AAAAAGCTCG CGCCAAAAAT ATTTGAATGTTTTGCGGGCG CGTTCGCCGG 551 CACGGTTTAC CGGTTTGTCT GCCTGTTCTA CATAATAAATGACGGAATCG 601 CCCATCATAT CTGCTCCTCA ACGTGTACGG TATCTGTTTG CACCTTACTG651 CGGCTTTCTG CCTTCGGCAT CCGATTCGGA TTTGAAAAGT TCCAAATATT 701 CGGAATAG

This encodes a protein having amino acid sequence <SEQ ID 554>:

1 MMKRRIAVFV LLMQKIRILG QLLPKIVNTV PAHRMLFQXF GMFFFFIHQQ 51 YLPGIAEIDSPCGIVFGTLL FRHXSTHCLY GKAAVGNAVA HEHPVADVVN 101 RNANAFALFD IGQFAGFIVQHAINVKTVKI NIVDPHMFAN FAXFAVLEKR 151 ALTMAKSKXX XMRRRSQKSS RQKYLNVLRARSPARFTGLS ACST**MTES 201 PIISAPQRVR YLFAPYCGFL PSASDSDLKS SKYSE*

ORF136a and ORF136-1 show 73.1% identity in 238 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF136 shows 92.3% identity over a 234aa overlap with a predicted ORF(ORF136ng) from N. gonorrhoeae:

The complete length ORF136ng nucleotide sequence <SEQ ID 555> is:

1 ATGATGAAGC GGCGTATAGC CGTCTTCGTC CTGCTCATGC AGAAAATCCG 51 GATTTTGGGACAACTGTTGC CGAAAATCGT CAATACAGTT CCGGCACATC 101 GGATGCTCTT CCAAATTTTCGGGATGTTCT TTTTCTTCAT ACACCGGCAA 151 TACCTGCCCG GGATCGCCGA AATCGATTCCCCAGGCGGTA TCGTGTTCGG 201 TACGCTCCTC TTCCGTCATC TGTCCGCGCA TTGCCTGTACGGTAAAGCCG 251 CCGTAGGGGA TGCCGTTGCA CACGAACATC CAGTCGCTGA TGTCGCCAAC301 CGGAACGCAA ACGCTTTCGC CTTGTTCGAC ATTGGTCAGT CCGCCGGGTT 351CATTGTTCAG CACACCGTAA ATATAAAGAC CGTCAAAATA AATATCGTCG 401 ATCCACATATGTTCGCAAAT TTCGCCGTCT TCGCCGTCTT GGAAAAAAGG 451 GACTTTGACC ATGGCAAAATCCAAGGCGGA AATAATGCGG CGGCGTTCCC 501 AAAAAAGCTC GCGCCAAAAG TATTTGAATGTTTTACGGGC GCGTTCGCCG 551 GCACGGTTTA CCGGTTCGTC TGCCTGTTCT ACATAATAAATGACGGAATC 601 GCCCATCATA CTGCTCCTCA ACGTGTACGG TATCTGTTTG CACCTTACCG651 CGGTTTTCTA CCTCCGGCAT CCGATTCGGA TTTGAAAAGT TCCAAATATT 701 CGGAATAG

This encodes a protein having amino acid sequence <SEQ ID 556>:

1 MMKRRIAVFV LLMQKIRILG QLLPKIVNTV PAHRMLFQIF GMFFFFIHRQ 51 YLPGIAEIDSPGGIVFGTLL FRHLSAHCLY GKAAVGDAVA HEHPVADVAN 101 RNANAFALFD IGQSAGFIVQHTVNIKTVKI NIVDPHMFAN FAVFAVLEKR 151 DFDHGKIQGG NNAAAFPKKLAPKVFECFTG AFAGTVYRFV CLFYIINDGI 201 AHHTAPQRVR YLFAPYRGFL PPASDSDLKSSKYSE*

ORF136ng and ORF136-1 show 93.6% identity in 235 aa overlap:

Based on the presence of the putative transmembrane domains in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 67

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 557>:

1 ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT TGGCAATCGC 51 CGCCGCCGCGTTGCTTGCCG CC.TGCGGAC GGCGGGAAAT AATGCTGTCC 101 GCAAGCCGGT GCAAACCGCCAAACCCGCCG CAGTGGTCGG TTTGGCACTC 151 GGTGGCGGCG CATCTAAAGG ATTTGCCCATGTAGGTATTA TTAAGGTTTT 201 GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACCTCCGCAGGTT 251 CGATTGTCGG CAACCTTTTT GCATCGGGTA TGTCGCCCGA CCGCCTCGAA301 TTGGAAGCCG AAATTTTAGG CAAAACCGAT TTGGTCGATT TAACCTTGTC 351CACCAATGGG TTTATCAAAG GCGCAAAGCT GCAAAATTAC ATCAACCGAA 401 AACTCCGCGGCATGCAGATT CAGCAGTTTC CCATCAAATT TGCCGCC..

This corresponds to the amino acid sequence <SEQ ID 558; ORF137>:

1 MENMVTFSKI RPLLAIAAAA LLAAXRTAGN NAVRKPVQTA KPAAVVGLAL 51 GGGASKGFAHVGIIKVLKEN GIPVKVVTGT SAGSIVGNLF ASGMSPDRLE 101 LEAEILGKTD LVDLTLSTNGFIKGAKLQNY INRKLRGMQI QQFPIKFAA..

Further work revealed the complete nucleotide sequence <SEQ ID 559>:

1 ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT TGGCAATCGC 51 CGCCGCCGCGTTGCTTGCCG CCTGCGGCAC GGCGGGAAAT AATGCTGTCC 101 GCAAGCCGGT GCAAACCGCCAAACCCGCCG CAGTGGTCGG TTTGGCACTC 151 GGTGGCGGCG CATCTAAAGG ATTTGCCCATGTAGGTATTA TTAAGGTTTT 201 GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACATCGGCAGGTT 251 CGATTGTCGG CAGCCTTTTT GCATCGGGTA TGTCGCCCGA CCGCCTCGAA301 TTGGAAGCCG AAATTTTAGG CAAAACCGAT TTGGTCGATT TAACCTTGTC 351CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC ATCAACCGAA 401 AAGTCGGCGGCAGGCAGATT CAGCAGTTTC CCATCAAATT TGCCGCCGTT 451 GCTACTGATT TTGAAACCGGCAAGGCCGTC GCTTTCAATC AGGGGAATGC 501 CGGGCAGGCT GTGCGCGCTT CCGCCGCCATTCCCAATGTG TTCCAACCCG 551 TTATCATCGG CAGGCATACA TATGTTGACG GCGGTCTGTCGCAGCCCGTG 601 CCCGTCAGTG CCGCCCGGCG GCAGGGGGCG AATTTCGTGA TTGCCGTCGA651 TATTTCCGCC CGTCCGGGCA AAAACATCAG CCAAGGTTTC TTCTCTTATC 701TCGATCAGAC GCTGAACGTA ATGAGCGTTT CTGCGTTGCA AAATGAGTTG 751 GGGCAGGCGGATGTGGTTAT CAAACCGCAG GTTTTGGATT TGGGTGCAGT 801 CGGCGGATTC GATCAGAAAAAACGCGCCAT CCGGTTGGGT GAGGAGGCAG 851 CACGTGCCGC ATTGCCTGAA ATCAAACGCAAACTGGCGGC ATACCGTTAT 901 TGA

This corresponds to the amino acid sequence <SEQ ID 560; ORF137-1>:

1 MENMVTFSKI RPLLAIAAAA LLAACGTAGN NAVRKPVQTA KPAAVVGLAL 51 GGGASKGFAHVGIIKVLKEN GIPVKVVTGT SAGSIVGSLF ASGMSPDRLE 101 LEAEILGKTD LVDLTLSTSGFIKGEKLQNY INRKVGGRQI QQFPIKFAAV 151 ATDFETGKAV AFNQGNAGQA VRASAAIPNVFQPVIIGRHT YVDGGLSQPV 201 PVSAARRQGA NFVIAVDISA RPGKNISQGF FSYLDQTLNVMSVSALQNEL 251 GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE IKRKLAAYRY301 *

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF137 shows 93.3% identity over a 149aa overlap with an ORF (ORF137a)from strain A of N. meningitidis:

The complete length ORF137a nucleotide sequence <SEQ ID 561> is:

1 ATGGAAAATA TGGTAACGTT TTCAAAAATC AGACCGCTTT TGGCAATCGC 51 CGCCGCCGCGTTGCTTGCCG CCTGCGGCAC GGCGGGAAAT AATGCTGCCC 101 GCAAGCCGGT GCAAACCGCCAAACCCGCCG CAGTGGTCGG TTTGGCACTC 151 GGTGGCGGCG CATCTAAAGG ATTTGCCCATGTAGGTATTA TTAAGGTTTT 201 GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACATCGGCAGGTT 251 CGATAGTCGG CAGCCTTTTT GCATCGGGTA TGTCGCCCGA CCGCCTCGAA301 TTGGAAGCCG AAATTTTAGG TAAAACCGAT TTGGTCGATT TAACCTTGTC 351CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC ATCAACCGAA 401 AAGTCGGCGGCAGGCGGATT CAGCAGTTTC CCATCAAATT TGCCGCCGTT 451 GCTACTGATT TTGAAACCGGCAAGGCCGTC GCTTTCAATC AAGGGAATGC 501 CGGGCAGGCT GTGCGCGCTT CCGCCGCCATTCCCAATGTG TTCCAACCCG 551 TTATCATCGG CAGGCATACA TATGTTGACG GCGGTCTGTCGCAGCCCGTG 601 CCCGTCAGTG CCGCCCGGCG GCANGNNNNG NATNTCGTGA TTGCCGTCGA651 TATTTCCGCC CGTCCGAGCA AAAACATCAG CCAAGGCTTC TTCTCTTATC 701TCGATCAGAC GCTGAACGTA ATGAGCGTTT CCGCGTTGCA AAATGAGTTG 751 GGGCAGGCGGATGTGGTTAT CAAACCGCAG GTTTTGGATT TGGGTGCAGT 801 CGGCGGATTC GATCAGAAAAAACGCGCCAT CCGGTTGGGT GAGGAGGCAG 851 CACGTGCCGC ATTGCCTGAA ATCAAACGCAAACTGGCGGC ATACCGTTAT 901 TGA

This encodes a protein having amino acid sequence <SEQ ID 562>:

1 MENMVTFSKI RPLLAIAAAA LLAACGTAGN NAARKPVQTA KPAAVVGLAL 51 GGGASKGFAHVGIIKVLKEN GIPVKVVTGT SAGSIVGSLF ASGMSPDRLE 101 LEAEILGKTD LVDLTLSTSGFIKGEKLQNY INRKVGGRRI QQFPIKFAAV 151 ATDFETGKAV AFNQGNAGQA VRASAAIPNVFQPVIIGRHT YVDGGLSQPV 201 PVSAARRXXX XXVIAVDISA RPSKNISQGF FSYLDQTLNVMSVSALQNEL 251 GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE IKRKLAAYRY301 *

ORF137a and ORF137-1 show 97.3% identity in 300 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF137 shows 89.9% identity over a 149aa overlap with a predicted ORF(ORF137ng) from N. gonorrhoeae:

The complete length ORF137ng nucleotide sequence <SEQ ID 563> is:

1 ATGGAAAATA TGGTAACGTT TTCAAAAATC AGATCATTTT TGGCAATCGC 51 CGCCGCCGCGTTGCTTGCCG CCTGCGGTAC GGCGGGAAAC AATGCCGCCC 101 GCAAGCCGGT GCAAACCGCCAAACCCGCCG CAGTGGTCGC TTTGGCACTC 151 GGTGGCGGCG CATCTAAAGG ATTTGCCCATATAGGAATTG TTAAGGTTTT 201 GAAAGAAAAC GGTATTCCTG TGAAGGTGGT TACCGGCACATCGGCAGGTT 251 CGATAGTCGG CAGCCTTTTG GCATCGGGTA TGTCGCCCGA CCGCCTCGAA301 TTGGAAGCCG AGATTTTAGG TAAAACCGAT TTAGTCGATT TAACCTTGTC 351CACCAGTGGT TTTATCAAAG GCGAAAAGCT GCAAAATTAC ATCAACCGAA 401 AAGTCGGCGGCAGGCAGATT CAGCAGTTTC CCATCAAATT TGCCGCCGTT 451 GCCACTGATT TTGAAACCGGCAAGGCCGTC GCTTTCAATC AAGGGAATGC 501 CGGGCAGGCG GTTCGTGCTT CCGCCGCCATTCCCAATGTG TTCCAGCCAG 551 TCATCATCGG CAGGCACAAA TATGTTGACG GCGGTCTGTCGCAGCCCGTG 601 CCCGTCAGTG CCGCTCGGCG GCAGGGGGCG AATTTCGTGA TTGCCGTCGA651 TATTTCCGCA CGTCCGAGCA AAAATGTCGG TCAAGGTTTC TTCTCTTATC 701TCGATCAGAC GCTGAACGTG ATGAGCGTTT CCGTGTTGCA AAACGAGTTG 751 gggcAGGCGGATGTGGTTAT CAAACCGCag gtTTTGGATT TGGGTGCAGT 801 CGGCGGATTC GATCAGAAAAAGCGCGCCAT CCGGTTGGGC GAGGAGGCAG 851 CACGTGCCGC ATTGCCTGAA ATCAAACGCAAACTGGCGGC ATACCGTTAT 901 TGA

This encodes a protein having amino acid sequence <SEQ ID 564>:

1 MENMVTFSKI RSFLAIAAAA LLAACGTAGN NAARKPVQTA KPAAVVALAL 51 GGGASKGFAHIGIVKVLKEN GIPVKVVTGT SAGSIVGSLL ASGMSPDRLE 101 LEAEILGKTD LVDLTLSTSGFIKGEKLQNY INRKVGGRQI QQFPIKFAAV 151 ATDFETGKAV AFNQGNAGQA VRASAAIPNVFQPVIIGRHK YVDGGLSQPV 201 PVSAARRQGA NFVIAVDISA RPSKNVGQGF FSYLDQTLNVMSVSVLQNEL 251 GQADVVIKPQ VLDLGAVGGF DQKKRAIRLG EEAARAALPE IKRKLAAYRY301 *

ORF137ng and ORF137-1 show 96.0% identity in 300 aa overlap:

Based on the presence of a predicted prokaryotic membrane lipoproteinlipid attachment site (underlined) in the gonococcal protein, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 68

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 565>:

1 ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA CCGCCATGCA 51CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGcTG CCGCTTTCCT 101GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT TTACCTTTTA 151AAGGAAGACC GCGCGCGCAT CGTCGCCmAT ATGCGGCAGG CGGGTTTGAA 201CCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG GCAAAAGGCG 251GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA CATAGAAACA 301ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG CTTTGGACAA 351ACACGAAGGG CTGCTATTC..

This corresponds to the amino acid sequence <SEQ ID 566; ORF138>:

1 MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN RLGHLAFYLL 51KEDRARIVAX MRQAGLNPDP KTVKAVFAET AKGGLELAPA FFRKPEDIET 101MFKAVHGWEH VQQALDKHEG LLF

Further work revealed the complete nucleotide sequence <SEQ ID 567>:

1 ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA CCGCCATGCA 51CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG CCGCTTTCCT 101GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT TTACCTTTTA 151AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGGCAGG CGGGTTTGAA 201CCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG GCAAAAGGCG 251GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA CATAGAAACA 301ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG CTTTGGACAA 351ACACGAAGGG CTGCTATTCA TCACGCCGCA CATCGGCAGC TACGATTTGG 401GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCCGCTGAC CGCCATGTAC 451AAACCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG CGGGCAGGGT 501TCGCGGCAAA GGAAAAACCG CGCCTACCAG CATACAAGGG GTCAAACAAA 551TCATCAAAGC CCTGCGTTCG GGCGAAGCAA CCATCGTCCT GCCCGACCAC 601GTCCCCTCCC CTCAAGAAGG CGGGGAAGGC GTATGGGTGG ATTTCTTCGG 651CAAACCTGCC TATACCATGA CGCTGGCGGC AAAATTGGCA CACGTCAAAG 701GCGTGAAAAC CCTGTTTTTC TGCTGCGAAC GCCTGCCTGG CGGACAAGGT 751TTCGATTTGC ACATCCGCCC CGTCCAAGGG GAATTGAACG GCGACAAAGC 801CCATGATGCC GCCGTGTTCA ACCGCAATGC CGAATATTGG ATACGCCGTT 851TTCCGACGCA GTATCTGTTT ATGTACAACC GCTACAAAAT GCCGTAA

This corresponds to the amino acid sequence <SEQ ID 568; ORF138-1>:

1 MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN RLGHLAFYLL 51KEDRARIVAN MRQAGLNPDP KTVKAVFAET AKGGLELAPA FFRKPEDIET 101MFKAVHGWEH VQQALDKHEG LLFITPHIGS YDLGGRYISQ QLPFPLTAMY 151KPPKIKAIDK IMQAGRVRGK GKTAPTSIQG VKQIIKALRS GEATIVLPDH 201VPSPQEGGEG VWVDFFGKPA YTMTLAAKLA HVKGVKTLFF CCERLPGGQG 251FDLHIRPVQG ELNGDKAHDA AVFNRNAEYW IRRFPTQYLF MYNRYKMP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF138 shows 99.2% identity over a 123aa overlap with an ORF (ORF138a)from strain A of N. meningitidis:

The complete length ORF138a nucleotide sequence <SEQ ID 569> is:

1 ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA CCGCCATGCA 51CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG CCGCTTTCCT 101GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT TTACCTTTTA 151AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGTCAGG CAGGCATGAA 201TCCCGACCCC AAAACGGTCA AAGCCGTTTT TGCGGAAACG GCAAAAGGCG 251GTTTGGAACT TGCCCCCGCG TTTTTCAGAA AACCGGAAGA CATAGAAACA 301ATGTTCAAAG CGGTACACGG CTGGGAACAT GTGCAGCAGG CTTTGGACAA 351ACACGAAGGG CTGCTATTCA TCACGCCGCA CATCGGCAGC TACGATTTGG 401GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCCGCTGAC CGCCATGTAC 451AAACCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG CGGGCAGGGT 501TCGCGGCAAA GGAAAAACCG CGCCTACCAG CATACAAGGG GTCAAACAAA 551TCATCAAAGC CCTGCGTTCG GGCGAAGCAA CCATCGTCCT GCCCGACCAC 601GTCCCCTCCC CTCAAGAAGG CGGGGAAGGC GTATGGGTGG ATTTCTTCGG 651CAAACCTGCC TATACCATGA CGCTGGCGGC AAAATTGGCA CACGTCAAAG 701GCGTGAAAAC CCTGTTTTTC TGCTGCGAAC GCCTGCCTGG CGGACAAGGT 751TTCGATTTGC ACATCCGCCC CGTCCAAGGG GAATTGAACG GCGACAAAGC 801CCATGATGCC GCCGTGTTCA ACCGCAATGC CGAATATTGG ATACGCCGTT 851TTCCGACGCA GTATCTGTTT ATGTACAACC GCTACAAAAT GCCGTAA

This encodes a protein having amino acid sequence <SEQ ID 570>:

  1 MFRLQFRLFP PLRTAMHILL TALLKCLSLL PLSCLHTLGN RLGHLAFYLL 51 KEDRARIVAN MRQAGLNPDP KTVKAVFAET AKGGLELAPA FFRKPEDIET101 MFKAVHGWEH VQQALDKHEG LLFITPHIGS YDLGGRYISQ QLPFPLTAMY151 KPPKIKAIDK IMQAGRVRGK GKTAPTSIQG VKQIIKALRS GEATIVLPDH201 VPSPQEGGEG VWVDFFGKPA YTMTLAAKLA HVKGVKTLFF CCERLPGGQG251 FDLHIRPVQG ELNGDKAHDA AVFNRNAEYW IRRFPTQYLF MYNRYKMP*

ORF138a and ORF138-1 show 99.7% identity over a 298aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF138 shows 94.3% identity over a 123aa overlap with a predicted ORF(ORF138ng) from N. gonorrhoeae:

The complete length ORF138ng nucleotide sequence <SEQ ID 571> is:

1 ATGTTTCGTT TACAATTCAG GCTGTTTCCC CCTTTGCGAA CCGCCATGCA 51CATCCTGTTG ACCGCCCTGC TCAAATGCCT CTCCCTGCTG TCGCTTTCCT 101GTCTGCACAC GCTGGGAAAC CGGCTCGGAC ATCTGGCGTT TTACCTTTTA 151AAGGAAGACC GCGCGCGCAT CGTCGCCAAT ATGCGGCAGG CGGGTTTGAA 201CCCCGACACG CAGACGGTCA AAGCCGTTTT TGCGGAAACG GCAAAATGCG 251GTTTGGAACT TGCCCCCGCG TTTTTCAAAA AACCGGAAGA CATCGAAACA 301ATGTTCAAAG CGGTACACGG CTGGGAACAC GTGCAGCAGG CTTTGGACAA 351GGGCGAAGGG CTGCTGTTCA TCACGCCGCA CATCGGCAGC TACGATTTGG 401GCGGACGCTA CATCAGCCAG CAGCTTCCGT TCCACCTGAC CGCCATGTAC 451AAGCCGCCGA AAATCAAAGC GATAGACAAA ATCATGCAGG CGGGCAGGGT 501GCGCGGCAAA GGCAAAACcg cgcccaccgg catACAAGGG GTCAAACAAA 551tcatcaAGGC CCTGCGCGCG GGCGAGGCAA CCAtcATCCT GCCCGACCAC 601GTCCCTTCTC CGCAGGAagg cggCGGCGTG TGGGCGGATT TTTTCGGCAA 651ACCTGCATAC acCATGACAC TGGCGGCAAA ATTGGCACAC GTCAAAGGCG 701TGAAAACCCT GTTTTTCTGC TGCGAACGCC TGCCCGACGG ACAAGGCTTC 751GTGTTGCACA TCCGCCCCGT CCAAGGGGAA TTGAACGGCA ACAAAGCCCA 801CGATGCCGCC GTGTTCAACC GCAATACCGA ATATTGGATA CGCCGTTTTC 851CGACGCAGTA TCTGTTTATG TACAACCGCT ATAAAACGCC GTAA

This encodes a protein having amino acid sequence <SEQ ID 572>:

1 MFRLQFRLFP PLRTAMHILL TALLKCLSLL SLSCLHTLGN RLGHLAFYLL 51KEDRARIVAN MRQAGLNPDT QTVKAVFAET AKCGLELAPA FFKKPEDIET 101MFKAVHGWEH VQQALDKGEG LLFITPHIGS YDLGGRYISQ QLPFHLTAMY 151KPPKIKAIDK IMQAGRVRGK GKTAPTGIQG VKQIIKALRA GEATIILPDH 201VPSPQEGGGV WADFFGKPAY TMTLAAKLAH VKGVKTLFFC CERLPDGQGF 251VLHIRPVQGE LNGNKAHDAA VFNRNTEYWI RRFPTQYLFM YNRYKTP*

ORF138ng and ORF138-1 show 94.3% identity over 299aa overlap:

In addition, ORF138ng is homologous to htrB protein from Pseudomonasfluorescens:

gnl|PID|e334283 (Y14568) htrB [Pseudomonas fluorescens] Length = 253Score = 80.8 bits (196), Expect = 9e−15 Identities =49/151 (32%), Positives = 79/151 (51%), Gaps = 6/151 (3%) Query: 101 MFKAVHGWEHVQQALDKGEGLLFITPHIGSYD-LGGRYISQQLPFHLTAMYKPPKIKAID 159 + +V G E +++AL  G+G++ IT H+G+++ L   Y SQ  P      Y+PPK+KA+D Sbjct: 94 LVREVEGLEVLKEALASGKGVVGITSHLGNWEVLNHFYCSQCKPI---IFYRPPKLKAVD 150 Query:160 KIMQAGRVRGKGKTAPTGIQGVKQIIKALRAGEATIILPDHVPSPQEGGGVWADFFGKPA 219++++  RV+   K A +  +G+  +IK +R G    I  D  P P E  G++  FF   A Sbjct: 151ELLRKQRVQLGNKVAASTKEGILSVIKEVRKGGQVGIPAD--PEPAESAGIFVPFFATQA 208 Query:220 YTMTLAAKLAHVKGVKTLFFCCERLPDGQGF 250  T      +        +F    RLPDG G+Sbjct: 209 LTSKFVPNMLAGGKAVGVFLHALRLPDGSGY 239

Based on this analysis, including the presence of a putativetransmembrane domain in the gonococcal protein, it was predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

ORF138-1 (57 kDa) was cloned in the pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 14A shows the results ofaffinity purification of the GST-fusion protein. Purified GST-fusionprotein was used to immunise mice, whose sera were used for ELISA(positive result) and FACS analysis (FIG. 14B). These experimentsconfirm that ORF138-1 is a surface-exposed protein, and that it is auseful immunogen.

Example 69

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 573>:

1 ..GCGTGGTCGG CCGGCGAATC GTGGCGTGTG TTAATGGAAA GTGAAACGTG 51  GCATGCGGTG TGGAATACTT TGCGCTTCTC GGCGGCGGCG GTGTATGCGG 101  CAGCGGTTTT GGGTGTGGTG TATGCGGCGC CGGCGCGGCG GTCGGCGTGG 151  ATGCGCGGGC TGATGTTTTA GCCGTTTATG GTGTCGCCGG TTTGTGTTTC 201  GGCGGGCGTG CTGCTGCTTT ATCCGCAGTG GACGGCTTCG TTGCCGTTGC 251  TGCTGGCGAT GTATGCGCTG CTGGCGTATC CGTTTGTGGC AAAAGATGTT 301  TTATCAGCCT GGGATGCACT GCCGCCGGAT TACGGCAGGG CGGCGGCGGG 351  TTTGGGTGCA AACGGCTTTC AGACGGCATG CCGCATCACG TTCCCCCTCT 401  TGAAACCGGC GTTGCGGCGC GGTCTGACTT TGGCGGCGGC AACCTGCGTG 451  GGCGAATTTG CGGCGACATT GTTTCTGTCG CGTCCGGAAT GGCAGACGCT 501  GACGACTTTG ATTTATGCCT ATTTGGGACG CGCGGGTGAG GATAATTACG 551  CGCGGGCGAT GGTGCTG..

This corresponds to the amino acid sequence <SEQ ID 574; ORF139>:

1 ..AWSAGESWRV LMESETWHAV WNTLRFSAAA VYAAAVLGVV YAAPARRSAW 51  MRGLMFXPFM VSPVCVSAGV LLLYPQWTAS LPLLLAMYAL LAYPFVAKDV 101  LSAWDALPPD YGRAAAGLGA NGFQTACRIT FPLLKPALRR GLTLAAATCV 151  GEFAATLFLS RPEWQTLTTL IYAYLGRAGE DNYARAMVL..

Further work revealed the complete nucleotide sequence <SEQ ID 575>:

1 ATGGATGGAC GGCGTTGGGT GGTATGGGGT GCTTTTGCCC TGCTGCCTTC 51GGCTTTTTTG GCGGTAATGG TCGTTGCGCC TTTGTGGGCG GTGGCGGCGT 101ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA TATGCTCAAA 151CGTTTGGCGT GGACGGTATT TCAGGCAGCG GCAACCTGTG TGCTGGTGCT 201GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG GCGTTTCCGG 251GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCTTTTGT GATGCCCACG 301TTGGTGGCGG GCGTGGGCGT GCTGGCCCTG TTCGGGGCGG ACGGGCTGTT 351GTGGCGCGGC AGGCAGGATA CGCCGTATCT GTTGTTGTAC GGCAATGTGT 401TTTTCAACCT TCCTGTGTTG GTCAGGGCGG CGTATCAGGG GTTTGTGCAA 451GTGCCTGCGG CACGGCTTCA GACGGCACGG ACGTTGGGCG CGGGGGCGTG 501GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG TGGCTTGCCG 551GCGGCGTGTG CCTTGTCTTT CTGTATTGTT TTTCCGGGTT CGGGCTGGCG 601CTGCTGCTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG AAATTTACCA 651GTTGGTCATG TTCGAACTCG ATATGGCGGT TGCTTCGGTG CTGGTGTGGC 701TGGTGTTGGG GGTAACGGCG GCGGCAGGGT TGCTGTATGC GTGGTTCGGC 751AGGCGCGCGG TTTCGGATAA GGCGGTTTCC CCTGTGATGC CGTCGCCGCC 801GCAGTCGGTC GGGGAATATG TGCTGCTGGC GTTTGCGGCG GCGGTGTTGT 851CTGTGTGCTG CCTGTTTCCT TTGTTGGCAA TTGTTGTGAA AGCGTGGTCG 901GCCGGCGAAT CGTGGCGTGT GTTAATGGAA AGTGAAACGT GGCAGGCGGT 951GTGGAATACT TTGCGCTTCT CGGCGGCGGC GGTGTATGCG GCGGCGGTTT 1001TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGTCGGCGTG GATGCGCGGG 1051CTGATGTTTT TGCCGTTTAT GGTGTCGCCG GTTTGTGTTT CGGCGGGCGT 1101GCTGCTGCTT TATCCGCAGT GGACGGCTTC GTTGCCGTTG CTGCTGGCGA 1151TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT TTTATCAGCC 1201TGGGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCGG GTTTGGGTGC 1251AAACGGCTTT CAGACGGCAT GCCGCATCAC GTTCCCCCTC TTGAAACCGG 1301CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CAACCTGCGT GGGCGAATTT 1351GCGGCGACAT TGTTTCTGTC GCGTCCGGAA TGGCAGACGC TGACGACTTT 1401GATTTATGCC TATTTGGGAC GCGCGGGTGA GGATAATTAC GCGCGGGCGA 1451TGGTGCTGAC ATTGCTGTTG GCGGCGTTCG CGCTGGGTAT TTTCCTGCTG 1501TTGGACGGCG GCGAAGGCGG AAAACAGACG GAAACGTTAT AA

This corresponds to the amino acid sequence <SEQ ID 576; ORF139-1>:

1 MDGRRWVVWG AFALLPSAFL AVMVVAPLWA VAAYDGLAWR AVLSDAYMLK 51RLAWTVFQAA ATCVLVLPLG VPVAWVLARL AFPGRALVLR LLMLPFVMPT 101LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL VRAAYQGFVQ 151VPAARLQTAR TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF LYCFSGFGLA 201LLLGGSRYAT VEVEIYQLVM FELDMAVASV LVWLVLGVTA AAGLLYAWFG 251RRAVSDKAVS PVMPSPPQSV GEYVLLAFAA AVLSVCCLFP LLAIVVKAWS 301AGESWRVLME SETWQAVWNT LRFSAAAVYA AAVLGVVYAA AARRSAWMRG 351LMFLPFMVSP VCVSAGVLLL YPQWTASLPL LLAMYALLAY PFVAKDVLSA 401WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT LAAATCVGEF 451AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAM VLTLLL AAFALGIFLL 501LDGGEGGKQT ETL*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF139 shows 94.7% identity over a 189aa overlap with an ORF (ORF139a)from strain A of N. meningitidis:

The complete length ORF139a nucleotide sequence <SEQ ID 577> is:

1 ATGGATGGAC GGCGTTGGGC GGTATGGGGT GCTTTTGCCC TGCTGCCTTC 51GGCTTTTTTG GCGGCAATGG TCGTTGCGCC TTTGTGGGCG GTGGCGGCGT 101ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA TATGCTCAAA 151CGTTTGGCGT GGACGGTATT TCAGGCAGCG GCAACCTGTG TGCTGGTGCT 201GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG GCGTTTCCGG 251GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCTTTTGT GATGCCCACG 301TTGGTGGCGG GCGTGGGCGT GCTGGCTCTG TTCGGGGCGG ACGGCCTGTN 351GTGGCGCGGC TGGCAGGATA CGCCGTATCT GTTGTTGTAC GGCAATGTGT 401TTTTTNACCT TCCTGTGTTG GTCAGGGCGG CATATCAGGG GTTTGTGCAA 451GTGCCTGCGG CACGGCTTCA GACGGCACNG ACATTGGGCG CGGGGGCGTG 501GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG TGGCTTGCCG 551GCGGCGTGTG CCTTGTCTTC CTGTATTGTT TTTCGGGGTT CGGGCTGGCA 601TTGCTGCTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG AAATTTACCA 651GTTGGTCATG TTCGAACTCG ATATGGCGGT TGCTTCGGTG CTNGTGTGGC 701TGGTGTNGGG GGTAACNGCG GCGGCAGGGT TGCTGTATGC GTGGTTCGGC 751AGGCGCGCGG TTTCGGATAA GGCNGTTTCC CCTGTGATGC CGTCGCCGCC 801GCAGTCGGTC GGGGAATATG TGCTNCTGGC GTTTGCGGCG GCGGTGTNGT 851CTGTGTGCTG CCTGTTTCNT TTGTTGGCAA TTGTTGTGAA AGCGTGGTCG 901GCCGGCGAAT CGTGGCGTGT GTTAATGGAA AGTGAAACGT GGCAGGCGGT 951GTGGAATACT NTGCGCTTCT CGGCGGCGGC GGTGTATGCG GCGGCGGTTT 1001TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGTCGGCGTG GATGCGCGGG 1051CTGATGTTTT TGCCGTTTAT GGTGTCGCCG GTTTGTGTTT CGGCGGGCGT 1101GCTGCTGCTT NATCCGCAGT GGACGGCTTC GTTGCCGCTG CTGCTGGCGA 1151TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT TTTATCAGCC 1201TGNGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCGG GTTTGGGTGC 1251AAACGGCTTT CAGACGGCAT GCCGCATCAC GTTCCCCCTC TTGAAACCGG 1301CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CAACCTGCGT GGGCGAATTT 1351GCGGCAACCT TGTTCNTGTC GCGTCNCGAG TGGCAGACGC TGACGACTTT 1401GATTTATGCC TATNTGGGAC GCGCGGGTGA NGATAATTAC GCGCGGGCGA 1451TGGTGCTGAC ATTGCTGTTG GCGGCGTTCG CGCTGGGTAT NTTCCTGCTG 1501TTGGACGGCG GCGAAGGCGG AAAACGGACG GAAACGTTAT AA

This encodes a protein having amino acid sequence <SEQ ID 578>:

1 MDGRRWAVWG AFALLPSAFL AAMVVAPLWA VAAYDGLAWR AVLSDAYMLK 51RLAWTVFQAA ATCVLVLPLG VPVAWVLARL AFPGRALVLR LLMLPFVMPT 101LVAGVGVLAL FGADGLXWRG WQDTPYLLLY GNVFFXLPVL VRAAYQGFVQ 151VPAARLQTAX TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF LYCFSGFGLA 201LLLGGSRYAT VEVEIYQLVM FELDMAVASV LVWLVXGVTA AAGLLYAWFG 251RRAVSDKAVS PVMPSPPQSV GEYVLLAFAA AVXSVCCLFX LLAIVVKAWS 301AGESWRVLME SETWQAVWNT XRFSAAAVYA AAVLGVVYAA AARRSAWMRG 351LMFLPFMVSP VCVSAGVLLL XPQWTASLPL LLAMYALLAY PFVAKDVLSA 401XDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT LAAATCVGEF 451AATLFXSRXE WQTLTTLIYA YXGRAGXDNY ARAMVLTLLL AAFALGXFLL 501LDGGEGGKRT ETL*

ORF139a and ORF139-1 show 96.5% homology over a 514aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF139 shows 95.2% identity over a 189aa overlap with a predicted ORF(ORF139ng) from N. gonorrhoeae:

The complete length ORF139ng nucleotide sequence <SEQ ID 579> ispredicted to encode a protein having amino acid sequence <SEQ ID 580>:

1 MDGRCWAVRG AFSLLPSAFL AVMVVAPLWA VAAYDGLAWR AVLSDAYMLK 51RLAWTVFQAA ATCVLVLPLG VPVAWVLARL AFPGRALVLR LLMLPFVMPT 101LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL VRAAYQGFAQ 151VPAARLQTAR TLGAGAWRPF WDIEMPVLRP WLAGGVCLVF LYCFSGFGLA 201LLLGGSRYAT VEVEIYQLVM FELDMAGASA LVWLVLGVTA AAGLLYAWFG 251RRAVSDKAVS PVMPSPPQSV GEYVLLAFSV AVLSVCCLFP LSAIVVKAWS 301AGESRRVLME SETWQAVWNT LRFSAAAVFA AAVLGVVYAA AARRLVWMRG 351LVFLPFMVSP VCVSAGVLLL YPGWTASLPL LLAMYALLAY PFVAKDVLSA 401WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT LAAATCVGEF 451AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAMVLTLLL SAFAVCIFLL 501LDNGEGGKRT ETL*

Further work revealed a variant gonococcal DNA sequence <SEQ ID 581>:

1 ATGGATGGAC GGTGTTGGGC GGTACGGGGT GCTTTTTCCC TGCTGCCTTC 51GGCTTTTTTG GCGGTAATGG TCGTTGCGCC TTTGTGGGCG GTGGCGGCGT 101ATGACGGTTT GGCGTGGCGC GCGGTGCTGT CGGATGCCTA TATGCTCAAA 151CGTTTGGCGT GGACGGTGTT TCAGGCGGCG GCAACCTGTG TGCTGGTGCT 201GCCTTTGGGC GTGCCTGTCG CGTGGGTGCT GGCGCGGCTG GCGTTCCCGG 251GGCGGGCTTT GGTGCTGCGC CTGCTGATGC TGCCGTTTGT GATGCCCACG 301CTGGTGGCGG GCGTGGGCGT GCTGGCTCTG TTCGGGGCGG ACGGGCTGTT 351GTGGCGCGGC CGGCAGGATA CGCCGTATCT GTTGTTGTAC GGCAATGTGT 401TTTTCAACCT GCCCGTGTTG GTCAGGGCGG CGTATCAGGG GTTTGCTCAA 451GTGCCTGCGG CACGGCTTCA GACGGCACGG ACGTTGGGCG CGGGGGCGTG 501GCGGCGGTTT TGGGACATTG AAATGCCCGT TTTGCGCCCG TGGCTTGCCG 551GCGGCGTGTG CCTTGTCTTC CTGTATTGTT TTTCGGGGTT CGGGCTGGCA 601TTGCTGTTGG GCGGCAGCCG TTATGCCACG GTCGAAGTGG AAATTTACCA 651GTTGGTTATG TTCGAACTCG ATATGGCGGG GGCTTCGGCG CTGGTGTGGC 701TGGTGTTGGG GGTAACGGCG GCGGCAGGGT TGCTGTATGC GTGGTTCGGC 751AGGCGCGCGG TTTCGGATAA GGCGGTTTCC CCCGTGATGC CGTCGCCGCC 801GCAATCGGTG GGGGAATATG TATTGCTGGC ATTTTCGGTG GCGGTGTTGT 851CCGTGTGCTG CCTGTTTCCT TTGTCGGCAA TTGTTGTGAA AGCGTGGTCG 901GCCGGCGAAT CGCGGCGTGT GTTAATGGAA AGTGAAACGT GGCAGGCAGT 951GTGGAATACt ttGCGCTTTT CGGCGGCGGC GGTGTTTGCG GCGGCGGTTT 1001TGGGTGTGGT GTATGCGGCG GCGGCGCGGC GGCTGGTGTG GATGCGCGGA 1051CTGGTGTTTT TACCGTTTAT GGTGTCGCCG GTTTGTGTTT CGGCGGGCGT 1101GCTGCTGCTT TATCCGGGGT GGACGGCTTC GTTACCGCTG CTGCTGGCGA 1151TGTATGCGCT GCTGGCGTAT CCGTTTGTGG CAAAAGATGT TTTATCGGCC 1201TGGGATGCAC TGCCGCCGGA TTACGGCAGG GCGGCGGCAG GTTTGGGCGC 1251AAACGGCTTT CAGACGGCAT GCCGTATCAC GTTCCCCCTC TTGAAACCGG 1301CGTTGCGGCG CGGTCTGACT TTGGCGGCGG CGACGTGTGT GGGCGAATTT 1351GCGGCAACCT TGTTCCTGTC GCGTCCGGAA TGGCAGACGT TGACGACTTT 1401GATTTATGCC TATTTGGGGC GTGCGGGTGA GGACAATTAT GCGCGGGCAA 1451TGGTGTTGAC ATTGCTGTTG TCGGCATTTG CGGTGTGCAT TTTCCTGCTG 1501TTGGACAACG GCGAAGGCGg aaaACGGACG GAAACGTTAT AA

This corresponds to the amino acid sequence <SEQ ID 582; ORF139ng-1>:

1 MDGRCWAVRG AFSLLPSAFL AVMVVAPLWA VAAYDGLAWR AVLSDAYMLK 51RLAWTVFQAA ATCVLVLPLG VPVAWVLARL AFPGRALVLR LLMLPFVMPT 101LVAGVGVLAL FGADGLLWRG RQDTPYLLLY GNVFFNLPVL VRAAYQGFAQ 151VPAARLQTAR TLGAGAWRRF WDIEMPVLRP WLAGGVCLVF LYCFSGFGLA 201LLLGGSRYAT VEVEIYQLVM FELDMAGASA LVWLVLGVTA AAGLLYAWFG 251RRAVSDKAVS PVMPSPPQSV GEYVLLAFSV AVLSVCCLFP LSAIVVKAWS 301AGESRRVLME SETWQAVWNT LRFSAAAVFA AAVLGVVYAA AARRLVWMRG 351LVFLPFMVSP VCVSAGVLLL YPGWTASLPL LLAMYALLAY PFVAKDVLSA 401WDALPPDYGR AAAGLGANGF QTACRITFPL LKPALRRGLT LAAATCVGEF 451AATLFLSRPE WQTLTTLIYA YLGRAGEDNY ARAMVLTLLL SAFAVCIFLL 501LDNGEGGKRT ETL*

ORF139ng-1 and ORF139-1 show 95.9% identity over 513aa overlap:

Based on the presence of a predicted binding-protein-dependent transportsystems inner membrane component signature (underlined) in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 70

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 583>:

1 ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT TGGGCATTTC 51GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAGA TTCCGCATCC 101ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC TTTGGCAACC 151GGTTTGCCCA CAGGCAGCAT TGTCAAAGAC ATACTGGTCA AAAACTTCGG 201CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC GCGATGCTCG 251AACGTTTGGT C...

This corresponds to the amino acid sequence <SEQ ID 584; ORF140>:

1 MDGWTQTLSA QTLLGISAAA IILILILIVR FRIHALLTLV IVSLLTALAT 51GLPTGSIVKD ILVKNFGGTL GGVALLVGLG AMLERLV..

Further work revealed the complete nucleotide sequence <SEQ ID 585>:

1 ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT TGGGCATTTC 51GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA TTCCGCATCC 101ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC TTTGGCAACC 151GGTTTGCCCA CAGGCAGCAT TGTCAACGAC ATACTGGTCA AAAACTTCGG 201CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC GCGATGCTCG 251GACGTTTGGT CGAAACATCC GGCGGCGCAC AGTCGCTGGC GGACGCGCTG 301ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCGCTGG GCGTTGCCTC 351GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA ATCGTCATGC 401TGCCCATCGT GTTCGCCACC GCACGGCGCA TGAAACAGGA CGTACTGCCC 451TTCGCGCTTG CCTCCATCGG CGCATTTTCC GTCATGCACG TCTTCCTGCC 501GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC GCGAACATCG 551GCCAAGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC ATGGTATTTC 601AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCACCATCC ATGTTCCCGT 651TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAACGACCTG CCGAAAGAAC 701CTGCCAAAGC AGGAACGGTC GTCGCCATCA TGCTGATTCC CATGCTGCTG 751ATTTTCCTGA ATACCGGCGT ATCGGCCCTC ATCAGCGAAA AACTCGTAAG 801TGCGGACGAA ACCTGGGTTC AGACGGCAAA AATAATCGGT TCGACACCGA 851TCGCCCTTCT GATTTCCGTA TTGGTCGCAC TGTTTGTCTT GGGACGCAAA 901CGCGGCGAAA GCGGCAGCGC GTTGGAAAAA ACCGTGGACG GCGCACTCGC 951CCCCGTCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT ATGTTCGGCG 1001GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA CAGCATGGCG 1051GATTTGGGCA TTCCCGTCCT TTTGGGCTGT TTCCTTGTCG CCTTGGCACT 1101GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACC GCCGCCGCGC 1151TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG GCAGCTCGCC 1201TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA GCCACTTCAA 1251CGACTCCGGC TTCTGGCTGG TCGGCCGTCT CTTGGACATG GACGTACCGA 1301CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC ACTCATCGGC 1351TTTGCCTTGT CCGCACTGCT GTTCGCCATC GTCTGA

This corresponds to the amino acid sequence <SEQ ID 586; ORF140-1>:

1 MDGWTQTLSA QTLLGISAAA IILILILIVK FRIHALLTLV IVSLLTALAT 51GLPTGSIVND ILVKNFGGTL GGVALLVGLG AMLGRLVETS GGAQSLADAL 101IRMFGEKRAP FALGVASLIF GFPIFFDAGL IVMLPIVFAT ARRMKQDVLP 151FALASIGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG LPTAFITWYF 201SGYMLGKVLG RTIHVPVPEL LSGGTQDNDL PKEPAKAGTV VAIMLIPMLL 251IFLNTGVSAL ISEKLVSADE TWVQTAKIIG STPIALLISV LVALFVLGRK 301RGESGSALEK TVDGALAPVC SVILITGAGG MFGGVLRASG IGKALADSMA 351DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA AAGFTDWQLA 401CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT VNQTLIALIG 451 FALSALLFAI V*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF140 shows 95.4% identity over a 87aa overlap with an ORF (ORF140a)from strain A of N. meningitidis:

The complete length ORF140a nucleotide sequence <SEQ ID 587> is:

1 ATGGACGGCT GGACACAGAC GCTGTCCGCG CAAACCCTGT TGGGCATTTC 51GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA TTCCGCATCC 101ACGCGCTGCT GACACTGGTC ATCGTCAGCC TGCTGACGGC TTTGGCAACC 151GGTTTGCCCA CAGGCAGCAT TGTCAACGAC GTACTGGTCA AAAACTTCGG 201CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGCCTGGGC GCGATGCTCG 251GACGTTTGGT CGAAACATCC GGCGGCGCAC AGTCGCTGGC GGACGCGCTG 301ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCGCTGG GCGTTGCCTC 351GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA ATCGTCATGC 401TGCCCATCGT GTTCGCCACC GCACGGCGCA TGAAACAGGA CGTACTGCCC 451TTCGCGCTTG CCTCCATCGG CGCATTTTCC GTCATGCACG TCTTCCTGCC 501GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC GCGAACATCG 551GCCAAGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC ATGGTATTTC 601AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCACCATCC ATGTTCCCGT 651TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAACGACCTG CCGAAAGAAC 701CTGCCAAAGC AGGAACGGTC GTCGCCATCA TGCTGATTCC CATGCTGCTG 751ATTTTCCTGA ATACCGGCGT ATCGGCCCTC ATCAGCGAAA AACTCGTAAG 801TGCGGACGAA ACCTGGGTTC AGACGGCAAA AATAATCGGT TCGACACCGA 851TCGCCCTTCT GATTTCCGTA TTGGTCGCAC TGTTTGTCTT GGGACGCAAA 901CGCGGCGAAA GCGGCAGCGC GTTGGAAAAA ACCGTGGACG GCGCACTCGC 951CCCCGTCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT ATGTTCGGCG 1001GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA CAGCATGGCG 1051GATTTGGGCA TTCCCGTCCT TTTGGGCTGT TTCCTTGTCG CCTTGGCACT 1101GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACC GCCGCCGCGC 1151TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG GCAGCTCGCC 1201TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA GCCACTTCAA 1251CGACTCCGGC TTCTGGCTGG TCGGCCGCCT CTTGGACATG GACGTACCGA 1301CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC ACTCATCGGC 1351TTTGCCTTGT CCGCACTGCT GTTCGCCATC GTCTGA

This encodes a protein having amino acid sequence <SEQ ID 588>:

1 MDGWTQTLSA QTLLGISAAA IILILILIVK FRIHALLTLV IVSLLTALAT 51GLPTGSIVND VLVKNFGGTL GGVALLVGLG AMLGRLVETS GGAQSLADAL 101IRMFGEKRAP FALGVASLIF GFPIFFDAGL IVMLPIVFAT ARRMKQDVLP 151FALASIGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG LPTAFITWYF 201SGYMLGKVLG RTIHVPVPEL LSGGTQDNDL PKEPAKAGTV VAIMLIPMLL 251IFLPNTGVSAL ISEKLVSADE TWVQTAKIIG STPIALLISV LVALFVLGRK 301RGESGSALEK TVDGALAPVC SVILITGAGG MFGGVLRASG IGKALADSMA 351DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA AAGFTDWQLA 401CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT VNQTLIALIG 451 FALSALLFAI V*

ORF140a and ORF140-1 show 99.8% identity over a 461aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF140 shows 92% identity over a 87aa overlap with a predicted ORF(ORF140ng) from N. gonorrhoeae:

The complete length ORF140ng nucleotide sequence <SEQ ID 589> waspredicted to encode a protein having amino acid sequence <SEQ ID 590>:

1 MDGRTQTLSA QTLLGISAAA IILILILIVK FRIRALLTLV IASLLTALAT 51GLPTGSIVND VLVKNFGGTL GGVALLVGLG AMLGRLVETS GGAQSLADAL 101IRMFGEKRAP FAPGVASLIF GFPIFFDAGL IVMLPIVFAT ARRMKQDVLP 151FALASVGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG LPTAFITWYF 201SGYMLGKVLG RAIHVPVPEL LSGGTQDSDP PKEPAKAGTV VAVMLIPMLL 251IFLNTGVSAL ISEKLVSADE TWVQTAKMIG STPVALLISV LAALLVLGRK 301RGESGSTLEK TVDGALAPAC SVILITGAGG MFGGVLRASG IGKALADSMA 351DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA AAGFTDWQLA 401CIVLATAAGS VGCSHFNDSG FWLVGRLSDM DVPTTLKTWT VNQTLIAFIG 451 FALSALLFAI V*

Further work revealed a variant gonococcal DNA sequence <SEQ ID 591>:

1 ATGGACGGCC GGACACAGAC GCTGTCCGCG CAAACCTTGT TGGGCATTTC 51GGCGGCGGCA ATCATCCTCA TTCTGATTTT AATCGTCAAA TTCCGCATCC 101GCGCGCTGCT GACACTGGTC ATCGCCAGCC TGCTGACGGC TTTGGCAACC 151GGTTTGCCCA CAGGCAGCAT CGTCAACGAC GTACTGGTCA AAAACTTCGG 201CGGCACGCTC GGCGGCGTGG CGCTTCTGGT CGGTCTGGGC GCAATGCTCG 251GACGTTTGGT AGAAACATCC GGCGGCGCAC AGTCGCTGGC GGACGCGCTG 301ATCCGGATGT TCGGCGAAAA ACGCGCACCG TTCGCTCCGG GCGTTGCCTC 351GCTGATTTTC GGCTTCCCGA TTTTCTTCGA TGCCGGACTA ATCGTCATGC 401TGCCCATCGT ATTCGCCACC GCACGGCGCA TGAAACAGGA CGTACTGCCC 451TTCGCGCTTG CCTCCGTCGG CGCATTTTCC GTCATGCACG TCTTCCTGCC 501GCCCCATCCG GGCCCGATTG CCGCTTCCGA ATTTTACGGC GCGAACATCG 551GCCAGGTTTT GATTTTGGGT CTGCCGACCG CCTTCATCAC ATGGTATTTC 601AGCGGCTATA TGCTCGGCAA AGTGTTGGGG CGCGCCATCC ATGTTCCCGT 651TCCCGAACTG CTCAGCGGCG GCACGCAAGA CAGCGACCCG CCGAAAGAAC 701CTGCCAAAGC AGGAACGGTC GTCGCCGTCA TGCTGATTCC CATGCTGCTG 751ATTTTCCTGA ATACCGGCGT ATCAGCCCTC ATCAGCGAAA AACTCGTAAG 801TGCGGACGAA ACTTGGGTTC AGACGGCAAA AATGATCGGT TCGACACCTG 851TCGCCCTTCT GATTTCCGTA TTGGCCGCAC TGTTGGTCTT GGGACGCAAA 901CGCGGCGAAA GCGGCAGCAC GTTGGAAAAA ACCGTGGACG GCGCACTCGC 951CCCCGCCTGT TCCGTGATTC TGATTACCGG CGCGGGCGGT ATGTTCGGCG 1001GCGTTTTGCG CGCTTCCGGC ATCGGCAAGG CACTCGCCGA CAGCATGGCG 1051GATTTGGGCA TTCCCGTCCT TTTGGGCTGC TTCCTTGTCG CCTTGGCACT 1101GCGTATCGCG CAAGGTTCGG CAACCGTCGC CCTGACCACA GCCGCCGCGC 1151TGATGGCTCC TGCCGTTGCC GCCGCCGGCT TTACCGACTG GCAGCTCGCC 1201TGTATCGTAT TGGCAACGGC GGCAGGTTCG GTCGGTTGCA GCCACTTCAA 1251CGACTCCGGC TTCTGGCTGG TCGGCCGCCT CTTGGATATG GACGTACCGA 1301CCACGCTGAA AACCTGGACG GTCAACCAAA CCCTCATCGC ATTCATCGGC 1351TTTGCCTTGT CCGCACTGCT GTTTGCCATC GTCTGA

This corresponds to the amino acid sequence <SEQ ID 592; ORF140ng-1>:

1 MDGRTQTLSA QTLLGISAAA IILILILIVK FRIRALLTLV IASLLTALAT 51GLPTGSIVND VLVKNFGGTL GGVALLVGLG AMLGRLVETS GGAQSLADAL 101IRMFGEKRAP FAPGVASLIF GFPIFFDAGL IVMLPIVFAT ARRMKQDVLP 151FALASVGAFS VMHVFLPPHP GPIAASEFYG ANIGQVLILG LPTAFITWYF 201SGYMLGKVLG RAIHVPVPEL LSGGTQDSDP PKEPAKAGTV VAVMLIPMLL 251IFLTGVSAL ISEKLVSADE TWVQTAKMIG STPVALLISV LAALLVLGRK 301RGESGSTLEK TVDGALAPAC SVILITGAGG MFGGVLRASG IGKALADSMA 351DLGIPVLLGC FLVALALRIA QGSATVALTT AAALMAPAVA AAGFTDWQLA 401CIVLATAAGS VGCSHFNDSG FWLVGRLLDM DVPTTLKTWT VNQTLIAFIG 451 FALSALLFAI V*

ORF140ng-1 and ORF140-1 show 96.3% identity over 461aa overlap:

Furthermore, ORF140ng-1 is homologous to an E. coli protein:

gi|882633 (U29579) ORF_o454 [Escherichia coli] >gi|1789097 (AE000358) o454;This 454 aa ORF is 34% identical (9 gaps) to 444 residues of an approx. 456 aa protein GNTP_BACLI SW: P46832 [Escherichia coli] Length = 454Score = 210 bits (529), Expect = 1e−53 Identities =130/384 (33%), Positives = 194/384 (49%), Gaps = 19/384 (4%) Query: 88ETSGGAQSLADALIRMFGEKRAPFAPGVASLIFGFPIFFDAGLIVMLPIVFATARRMKQD 147E SGGA+SLA+   R  G+KR   A  +A+   G P+FFD G I++ PI++  A+  K Sbjct: 80EHSGGAESLANYFSRKLGDKRTIAALTLAAFFLGIPVFFDVGFIILAPIIYGFAKVAKIS 139 Query:148 VLPFALASVGAFSVMHVFLPPHPGPIAASEFYGANIGQVLILGLPTAFITWYFSGYMLGK 207 L F L   G    +HV +PPHPGP+AA+    A+IG + I+G+  + I    GY   K Sbjct: 140PLKFGLPVAGIMLTVHVAVPPHPGPVAAAGLLHADIGWLTIIGIAIS-IPVGVVGYFAAK 198 Query:208 VLGRAIHVPVPELL----------SGGTQDSDPPKEPAKAGTVVAVMLIPMLLIFLNTGV 257++ +  +    E+L           G T+ SD    P  A  V ++++IP+ +I   T Sbjct: 199IINKRQYAMSVEVLEQMQLAPASEEGATKLSDKINPPGVA-LVTSLIVIPIAIIMAGT-- 255 Query:258 SALISEKLVSADETWVQTAKMIGSTPXXXXXXXXXXXXXXGRKRGESGSTLEKTVDGALA 317   +S  L+      + T ++IGS                  +RG S       +  AL Sbjct: 256---VSATLMPPSHPLLGTLQLIGSPMVALMIALVLAFWLLALRRGWSLQHTSDIMGSALP 312 Query:318 PACSVILITGAGGMFGGVLRASGIGKALADSMADLGIPVLLGCFLVALALRIAQGSXXXX 377 A  VIL+TGAGG+FG VL  SG+GKALA+ +  + +P+L   F+++LALR +QGS Sbjct: 313TAAVVILVTGAGGVFGKVLVESGVGKALANMLQMIDLPLLPAAFIISLALRASQGS--AT 370 Query:378 XXXXXXXXXXXXXXXGFTDWQLACIVLATAAGSVGCSHFNDSGFWLVGRLLDMDVPTTLK 437               G   Q   + LA   G +G SH NDSGFW+V + L + V   LK Sbjct: 371VAILTTGGLLSEAVMGLNPIQCVLVTLAACFGGLGASHINDSGFWIVTKYLGLSVADGLK 430 Query:438 TWTVNQTLIAFIGFALSALLFAIV 461 TWTV  T++ F GF ++  ++A++ Sbjct: 431TWTVLTTILGFTGFLITWCVWAVI 454

Based on this analysis, including the identification of the presence ofa putative leader sequence (double-underlined) and several putativetransmembrane domains (single-underlined) in the gonococcal protein, itis predicted that the proteins from N. meningitidis and N. gonorrhoeae,and their epitopes, could be useful antigens for vaccines ordiagnostics, or for raising antibodies.

Example 71

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 593>:

1 ..GATTTCGGCA TATCGCCCGT GTATCTTTGG GTTGCCGCCG CGTTCAAACA 51  TTTGCTGTCG CCGTGGGCTG CCGACTCATA CGATGTCGCA CGCTTTGCAG 101  GCGTATTTTT TGCCGTTATC GGACTGACTT CCTGCGGCTT TGCCGGTTTC 151  AACTTTTTGG GCAGACACCA CGGGCGCAC. GTCGTCCTGA TTCTCATCGG 201  CTGTATCGGG CTGATTCCAG TTGCCCATTT CCTCAACCCC GCTGCCGCCG 251  CCTTTGCCGC CGCCGGACTG GTGCTGCACG GTTATTCTTT GGCTCGCCGG 301  CGCGTGATTG CCGCCTCTTT TCTGCTCGGT ACGGGCTGGA CGCTGATGTC 351  GTTGGCAGCA GCTTATCCGG CAGCATTTGC CCTGATGCTG CCCTTGCCCG 401  TACTGATGTT TTTCCGTCCG ..

This corresponds to the amino acid sequence <SEQ ID 594; ORF141>:

1 ..DFGISPVYLW VAAAFKHLLS PWAADSYDVA RFAGVFFAVI GLTSCGFAGF 51  NFLGRHHGRX VVLILIGCIG LIPVAHFLNP AAAAFAAAGL VLHGYSLARR 101  RVIAASFLLG TGWTLMSLAA AYPAAFALML PLPVLMFFRP ..

Further work revealed the complete nucleotide sequence <SEQ ID 595>:

1 ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA AAACCCACGA 51AAAGCCGTGG CTGCTGCTGT TGATGGCGTT TGCCTGGTTG TGGCCCGGCG 101TGTTTTCCCA CGATTTGTGG AATCCTGACG AACCTGCCGT CTATACCGCC 151GTCGAAGCAC TGGCAGGCAG CCCCACCCCC TTGGTTGCCC ATCTGTTCGG 201TCAAACCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT GCCGCCGCGT 251TCAAACATTT GCTGTCGCCG TGGGCTGCCG ACTCATACGA TGCCGCACGC 301TTTGCAGGCG TATTTTTTGC CGTTATCGGA CTGACTTCCT GCGGCTTTGC 351CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAgCGTC GTCCTGATTC 401TCATCGGCTG TATCGGGCTG ATTCCAGTTG CCCATTTCCT CAACCCCGCT 451GCCGCCGCCT TTGCCGCCGC CGGACTGGTG CTGCACGGTT ATTCTTTGGC 501TCGCCGGCGC GTGATTGCCG CCTCTTTTCT GCTCGGTACG GGCTGGACGC 551TGATGTCGTT GGCAGCAGCT TATCCGGCAG CATTTGCCCT GATGCTGCCC 601TTGCCCGTAC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC GTTTGATGTT 651GACGGCAGTC GCCTCACTTG CCTTTGCCCT GCCGCTTATG ACCGTTTACC 701CGCTGCTCTT GGCAAAAACG CAGCCCGCGC TGTTCGCGCA ATGGCTCGAC 751TATCACGTTT TCGGTACGTT CGGCGGCGTG CGGCACGTTC AGACGGCATT 801CAGTTTGTTT TACTATCTGA AAAACCTGCT TTGGTTTGCA TTGCCCGCGC 851TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CGCGCCTGTT TTCGACCGAC 901TGGGGGATTT TGGGCGTCGT CTGGATGCTT GCCGTTTTGG TGCTGCTTGC 951CGTCAATCCG CAGCGTTTTC AGGATAACCT CGTCTGGCTG CTTCCGCCGC 1001TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGGCG CGGCGCGGCG 1051GCGTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGACTGT TTGCCGTGTT 1101CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC GCCAAGCTTG 1151CCGAACGCGC CGCCTATTTC AGCCCGTATT ATGTTCCTGA TATCGATCCC 1201ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC TGTGGGCGAT 1251TACCCGGAAA AACATACGCG GCAGGCAGGC GGTTACCAAC TGGGCGGCAG 1301GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT GCCGTGGCTG 1351GACGCGGCGA AAAGCCACGC GCCGGTCGTC CGGAGTATGG AGGCATCGCT 1401TTCCCCGGAA TTGAAACGGG AGCTTTCAGA CGGCATCGAG TGTATCGGCA 1451TAGGCGGCGG CGACCTGCAC ACGCGGATTG TTTGGACGCA GTACGGCACA 1501TTGCCGCACC GCGTCGGCGA TGTACAATGC CGCTACCGCA TCGTCCTCCT 1551GCCCCAAAAT GCGGATGCGC CGCAAGGCTG GCAGACGGTT TGGCAGGGTG 1601CGCGTCCGCG CAACAAAGAC AGTAAGTTCG CACTGATACG GAAAATCGGG 1651GAAAATATAT AA

This corresponds to the amino acid sequence <SEQ ID 596; ORF141-1>:

  1 MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW NPDEPAVYTA  51VEALAGSPTP LVAHLFGQTD FGIPPVYLWV AAAFKHLLSP WAADSYDAAR 101FAGVFFAVIG LTSCGFAGFN FLGRHHGRSV VLILIGCIGL IPVAHFLNPA 151AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA YPAAFALMLP 201LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TVYPLLLAKT QPALFAQWLD 251YHVFGTFGGV RHVQTAFSLF YYLKNLLWFA LPALPLAVWT VCRTRLFSTD 301WGILGVVWML AVLVLLAVNP QRFQDNLVWL LPPLALFGAA QLDSLRRGAA 351AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF SPYYVPDIDP 401IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA LLMTLFLPWL 451DAAKSHAPVV RSMEASLSPE LKRELSDGIE CIGIGGGDLH TRIVWTQYGT 501LPHRVGDVQC RYRIVLLPQN ADAPQGWQTV WQGARPRNKD SKFALIRKIG 551 ENI*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF141 shows 95.0% identity over a 140aa overlap with an ORF (ORF141a)from strain A of N. meningitidis:

The complete length ORF141a nucleotide sequence <SEQ ID 597> is:

1 ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA AAACCCACGA 51AAAGCCGTGG CTGTTGCTGT TGATGGCGTT TGCCTGGTTG TGGCCCGGCG 101TGTTTTCCCA CGATTTGTGG AATCCTGACG AACCTGCCGT CTATACCGCC 151GTCGAAGCAC TGGCAGGCAG CCCCACCCCT TTGGTTGCCC ATCTGTTCGG 201TCAAATCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT GCCGCCGCGT 251TCAAACATTT GCTGTCGCCG TGGGCTGCCG ACCCGTATGA TGCCGCACGC 301TTTGCCGGCG TGTTTTTCGC CGTTGTCGGA CTGACTTCCT GCGGCTTTGC 351CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAGCGTC GTCCTGATTC 401TCATCGGCTG TATCGGGCTG ATTCCGACCG TACACTTTCT CAACCCCGCT 451GCCGCCGCCT TTGCCGCCGC CGGACTGGTG CTGCACGGTT ATTCTTTGGC 501TCGCCGGCGC GTGATTGCCG CCTCTTTTCT GCTCGGTACG GGTTGGACGC 551TGATGTCGTT GGCAGCAGCT TATCCGGCGG CATTTGCCCT GATGCTGCCC 601CTGCCCGTGC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC GTTTGATGTT 651GACGGCAGTC GCCTCGCTTG CCTTTGCCCT GCCGCTTATG ACCGTTTACC 701CGCTGCTCTT GGCAAAAACG CAGCCCGCGC TGTTCGCGCA ATGGCTCGAC 751GATCACGTTT TCGGTACGTT CGGCGGCGTG CGGCACATTC AGACGGCATT 801CAGTTTGTTT TACTATCTGA AAAACCTGCT TTGGTTTGCA TTGCCTGCGC 851TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CGCGCCTGTT TTCGACCGAC 901TGGGGGATTT TGGGCGTCGT CTGGATGCTT GCCGTTTTGG TGCTGCTTGC 951CGTCAATCCG CAGCGTTTTC AGGATAACCT CGTCTGGCTG CTTCCGCCGC 1001TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGACG CGGCGCGGCG 1051GCGTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGACTGT TTGCCGTGTT 1101CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC GCCAAGCTTG 1151CCGAACGCGC CGCCTATTTC AGCCCGTATT ATGTTCCTGA TATCGATCCC 1201ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC TGTGGGCGAT 1251TACCCGCAAA AACATACGCG GCAGGCAGGC GGTTACCAAC TGGGCGGCAG 1301GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT GCCGTGGCTG 1351GACGCGGCGA AAAGCCACGC GCCCGTCGTC CGGAGTATGG AGGCATCGCT 1401TTCCCCGGAA TTAAAACGGG AGCTTTCAGA CGGCATCGAG TGTATCGACA 1451TAGGCGGCGG CGACCTACAC ACGCGGATTG TTTGGACGCA GTACGGCACA 1501TTGCCGCACC GCGTCGGCGA TGTACAATGC CGCTACCGCA TCGTCCGCTT 1551GCCCCAAAAC GCGGATGCGC CGCAAGGCTG GCAGACGGTC TGGCAGGGTG 1601CGCGCCCGCG CAACAAAGAC AGTAAGTTCG CACTGATACG GAAAACCGGG 1651GAAAATATAT TAAAAACAAC AGATTGA

This encodes a protein having amino acid sequence <SEQ ID 598>:

  1 MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW NPDEPAVYTA  51VEALAGSPTP LVAHLFGQID FGIPPVYLWV AAAFKHLLSP WAADPYDAAR 101FAGVFFAVVG LTSCGFAGFN FLGRHHGRSV VLILIGCIGL IPTVHFLNPA 151AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA YPAAFALMLP 201LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TVYPLLLAKT QPALFAQWLD 251DHVFGTFGGV RHIQTAFSLF YYLKNLLWFA LPALPLAVWT VCRTRLFSTD 301WGILGVVWML AVLVLLAVNP QRFQDNLVWL LPPLALFGAA QLDSLRRGAA 351AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF SPYYVPDIDP 401IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA LLMTLFLPWL 451DAAKSHAPVV RSMEASLSPE LKRELSDGIE CIDIGGGDLH TRIVWTQYGT 501LPHRVGDVQC RYRIVRLPQN ADAPQGWQTV WQGARPRNKD SKFALIRKTG 551 ENILKTTD*

ORF141a and ORF141-1 show 98.2% identity in 553 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF141 shows 95% identity over a 140aa overlap with a predicted ORF(ORF141ng) from N. gonorrhoeae:

An ORF141ng nucleotide sequence <SEQ ID 599> was predicted to encode aprotein having amino acid sequence <SEQ ID 600>:

  1 MPSEAVSARP LCEYLLHLAI RPFLLTLMLT YTPPDARPPA KTHEKPWLLL  51LMAFAWLWPG VFSHDLWNPA EPAVYTAVEA LAGSPTPLVA HLFGQTDFGI 101PPVYLWVAAA FKHLLSPWAA HPYDAARFAG VFFAVIGLTS CGFAGFNFLG 151RHHGRSVVLI HIGCIGLIPV AHFFNPAAAA FAAAGLVLHG YSLARRRVIA 201ASFLLGTGWT LMSLAAAYPA AFALMLPLPV LMFFRPWQSR RLMLTAVASL 251AFALPLMTVY PLLLAKTQPA LFAQWLNYHV FGTFGGVRHI QRAFSLFHYL 301KNLLWFAPPG LPLAVWTVCR TRLFSTDWGI LGIVWMLAVL VLLAFNPQRF 351QDNLVWLLPP LALFGAAQLD SLRRGAAAFV NWFGIMAFGL FAVFLWTGFF 401AMNYGWPAKL AERAAYFSPY YVPDIDPIPM AVAVLFTPLW LWAITRKNIR 451GRQAVTNWAA GVTLTWALLM TLFLPWLDAA KSHAPVVRSM EASFSPELKR 501ELSDGIECIG IGGGDLHTRI VWTQYGTLPH RVGDVRCRYR IVRLPQNADA 551PQGWQTVWQG ARPRNKDSKF ALIRKIGENI LKTTD*

Further work revealed the following gonococcal DNA sequence <SEQ ID601>:

1 ATGCTGACCT ATACCCCGCC CGATGCCCGC CCGCCCGCCA AAACCCACGA 51AAAACCGTGG CTGCTGCTGT TGATGGCGTT TGCCTGGCTG TGGCCCGGCG 101TGTTTTCCCA CGATTTGTGG AATCCTGCCG AACCTGCCGT CTATACCGCC 151GTCGAAGCAC TGGCAGGCAG CCCCACCCCC TTGGTTGCCC ATCTGTTCGG 201TCAAACCGAT TTCGGCATAC CGCCCGTGTA TCTTTGGGTT GCCGCCGCAT 251TCAAACATTT GCTGTCGCCG TGGGCAGCCG ACCCGTATGA TGCCGCACGC 301TTTGCAGGCG TATTTTTTGC CGTTATCGGA CTGACTTCTT GCGGCTTTGC 351CGGTTTCAAC TTTTTGGGCA GACACCACGG GCGCAGCGTT GTTTTAATCC 401ATATCGGCTG TATCGGGCTG ATTCCGGTTG CCCATTTCCT CAATCCcgcc 451gccgccgcct tTGCCGCCGC CGGACTGGTG CTGCacggct actcgctgGC 501ACGCCGGCGC GTGATtgccg cctctTtccT GCTCGGTACG GGTTGGACGT 551TGATGTCGCT GGCGGCAGCT TATCCGGCGG CGTTTGCGCT GATGCTGCCC 601CTGCCCGTGC TGATGTTTTT CCGTCCGTGG CAAAGCAGGC GTTTGATGTT 651GACGGCAGTC GCCTCGCTTG CCTTTGCCCT GCCGCTTATG ACCGTTTACC 701CGCTGCTCtt gGCAAAAACG CAGCCCGCGC TGTTTGCGCA ATGGCTCAAC 751TATCACGTTT TCGGTACGTt cggcgGCGTG CGGCAcaTTC AGAggGCatT 801Cagtttgttt cactatctgA AAaatctgct ttggttcgca ccgcccgggC 851TGCCGCTGGC GGTTTGGACG GTTTGCCGCA CACGCCTGTT TTCGACCGAC 901TGGGGGATTT TGGGCATTGT CTGGATGCTT GCCGTTTTGG TGCTGCTCGC 951CTTTAATCCG CAGCGTTTTC AAGACAACCT CGTCTGGCTG CTGCCGCCGC 1001TTGCCCTGTT CGGCGCGGCG CAACTGGACA GCCTGAGGCG CGGCGCGGCG 1051GCTTTTGTCA ACTGGTTCGG CATTATGGCG TTCGGGCTGT TTGCCGTGTT 1101CCTGTGGACG GGCTTTTTCG CCATGAATTA CGGCTGGCCC GCCAAGCTTG 1151CCGAACGCGC CGCCTACTTC AGCCCGTATT ACGTTCCCGA CATCGATCCC 1201ATTCCGATGG CGGTTGCCGT ACTGTTCACA CCCTTGTGGC TGTGGGCGAT 1251TACCCGGAAA AACATACGCG GCAGGCAGGC GGTTACCAAC TGGGCGGCAG 1301GCGTTACCCT GACCTGGGCT TTGCTGATGA CGCTGTTCCT GCCGTGGCTG 1351GACGCGGCGA AAAGCCACGC GCCCGTCGTC CGGAGTATGG AGGCATCGTT 1401TTCCCCGGAA TTAAAACGGG AGCTTTCAGA CGGCATCGAG TGTATCGGCA 1451TAGGCGGCGG CGACCTGCAC ACGCGGATTG TTTGGACGCA GTACGGCACA 1501TTGCCGCACC GCGTCGGCGA TGTCCGTTGC CGCTACCGTA TCGTCCGCCT 1551GCCCCAAAAC GCGGATGCGC CGCAAGGCTG GCAGACGGTC TGGCAGGGTG 1601CGCGCCCGCG CAACAAAGAC AGTAAGTTTG CACTGATACG GAAAATCGGG 1651GAAAATATAT TAAAAACAAC AGATTGA

This corresponds to the amino acid sequence <SEQ ID 602; ORF141ng-1>:

  1 MLTYTPPDAR PPAKTHEKPW LLLLMAFAWL WPGVFSHDLW NPAEPAVYTA  51VEALAGSPTP LVAHLFGQTD FGIPPVYLWV AAAFKHLLSP WAADPYDAAR 101FAGVFFAVIG LTSCGFAGFN FLGRHHGRSV VLIHIGCIGL IPVAHFLNPA 151AAAFAAAGLV LHGYSLARRR VIAASFLLGT GWTLMSLAAA YPAAFALMLP 201LPVLMFFRPW QSRRLMLTAV ASLAFALPLM TVYPLLLAKT QPALFAQWLN 251YHVFGTFGGV RHIQRAFSLF HYLKNLLWFA PPGLPLAVWT VCRTRLFSTD 301WGILGIVWML AVLVLLAFNP QRFQDNLVWL LPPLALFGAA QLDSLRRGAA 351AFVNWFGIMA FGLFAVFLWT GFFAMNYGWP AKLAERAAYF SPYYVPDIDP 401IPMAVAVLFT PLWLWAITRK NIRGRQAVTN WAAGVTLTWA LLMTLFLPWL 451DAAKSHAPVV RSMEASFSPE LKRELSDGIE CIGIGGGDLH TRIVWTQYGT 501LPHRVGDVRC RYRIVRLPQN ADAPQGWQTV WQGARPRNKD SKFALIRKIG 551 ENILKTTD*

ORF141ng-1 and ORF141-1 show 97.5% identity in 553 aa overlap:

Based on the presence of several putative transmembrane domains in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 72

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 603>:

  1 ..CAATCCGCCA AATGGTTATC GGGCCAAACT CTAGTCGGCA CAGCAATTGG  51  GATACGCGGG CAGATAAAGC TTGGCGGCAA CCTGCATTAC GATATATTTA 101  CCGGCCGCGC ATTGAAAAAG CCCGAATTTT TCCAATCAAG GAAATGGGCA 151  AGCGGTTTTC AGGTAGGCTA TACGTTTTAA

This corresponds to the amino acid sequence <SEQ ID 604; ORF142>:

 1 ..QSAKWLSGQT LVGTAIGIRG QIKLGGNLHY DIFTGRALKK PEFFQSRKWA 51  SGFQVGYTF*

Further work revealed the complete nucleotide sequence <SEQ ID 605>:

  1 ATGGATAATT CGGGTAGTGA GGCGACAGGA AAATACCAAG GAAATATCAC  51TTTCTCTGCC GACAATCCTT TGGGACTGAG TGATATGTTC TATGTAAATT 101ATGGACGTTC GATTGGCGGT ACGCCCGATG AGGAAAGTTT TGACGGCCAT 151CGCAAAGAAG GCGGATCAAA CAATTACGCC GTACATTATT CAGCCCCTTT 201CGGTAAATGG ACATGGGCAT TCAATCACAA TGGCTACCGT TACCATCAGG 251CAGTTTCCGG ATTATCGGAA GTCTATGACT ATAATGGAAA AAGTTACAAT 301ACTGATTTCG GCTTCAACCG CCTGTTGTAT CGTGATGCCA AACGCAAAAC 351CTATCTCGGT GTAAAACTGT GGATGAGGGA AACAAAAAGT TACATTGATG 401ATGCCGAACT GACTGTACAA CGGCGTAAAA CTGCGGGTTG GTTGGCAGAA 451CTTTCCCACA AAGAATATAT CGGTCGCAGT ACGGCAGATT TTAAGTTGAA 501ATATAAACGC GGCACCGGCA TGAAAGATGC TCTGCGCGCG CCTGAAGAAG 551CCTTTGGCGA AGGCACGTCA CGTATGAAAA TTTGGACGGC ATCGGCTGAT 601GTAAATACTC CTTTTCAAAT CGGTAAACAG CTATTTGCCT ATGACACATC 651CGTTCATGCA CAATGGAACA AAACCCCGCT AACATCGCAA GACAAACTGG 701CTATCGGCGG ACACCACACC GTACGTGGCT TCGACGGTGA AATGAGTTTG 751TCTGCCGAGC GGGGATGGTA TTGGCGCAAC GATTTGAGCT GGCAATTTAA 801ACCAGGCCAT CAGCTTTATC TTGGGGCTGA TGTAGGACAT GTTTCAGGAC 851AATCCGCCAA ATGGTTATCG GGCCAAACTC TAGTCGGCAC AGCAATTGGG 901ATACGCGGGC AGATAAAGCT TGGCGGCAAC CTGCATTACG ATATATTTAC 951CGGCCGCGCA TTGAAAAAGC CCGAATTTTT CCAATCAAGG AAATGGGCAA 1001 GCGGTTTTCA GGTAGGCTAT ACGTTTTAA

This corresponds to the amino acid sequence <SEQ ID 606; ORF142-1>:

  1 MDNSGSEATG KYQGNITFSA DNPLGLSDMF YVNYGRSIGG TPDEESFDGH  51RKEGGSNNYA VHYSAPFGKW TWAFNHNGYR YHQAVSGLSE VYDYNGKSYN 101TDFGFNRLLY RDAKRKTYLG VKLWMRETKS YIDDAELTVQ RRKTAGWLAE 151LSHKEYIGRS TADFKLKYKR GTGMKDALRA PEEAFGEGTS RMKIWTASAD 201VNTPFQIGKQ LFAYDTSVHA QWNKTPLTSQ DKLAIGGHHT VRGFDGEMSL 251SAERGWYWRN DLSWQFKPGH QLYLGADVGH VSGQSAKWLS GQTLVGTAIG 301IRGQIKLGGN LHYDIFTGRA LKKPEFFQSR KWASGFQVGY TF*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. gonorrhoeae

ORF142 shows 88.1% identity over a 59aa overlap with a predicted ORF(ORF142ng) from N. gonorrhoeae:

The complete length ORF142ng nucleotide sequence <SEQ ID 607> is:

  1 ATGGATAATT CGGGTAGTGA GGCGACAGGA AAATACCAAG GAAATATCAC  51TTTCTCTGCC GACAATCCTT TTGGACTGAG TGATATGTTC TATGTAAATT 101ATGGACGTTC AATTGGCGGT ACGCCCGATG AGGAAAATTT TGACGGCCAT 151CGCAAAGAAG GCGGATCAAA CAATTACGCC GTACATTATT CAGCCCCTTT 201CGGTAAATGG ACATGGGCAT TCAATCACAA TGGCTACCGT TACCATCAGG 251CGGTTTCCGG ATTATCGGAA GTCTATGACT ATAATGGAAA AAGTTACAAC 301ACTGATTTCG GCTTCAACCG CCTGTTGTAT CGTGATGCCA AACGCAAAAC 351CTATCTCAGT GTAAAACTGT GGACGAGGGA AACAAAAAGT TACATTGATG 401ATGCCGAACT GACTGTACAA CGGCGTAAAA CCACAGGTTG GTTGGCAGAA 451CTTTCCCACA AAGGATATAT CGGTCGCAGT ACGGCAGATT TTAAGTTGAA 501ATATAAACAC GGCACCGGCA TGAAAGATGC TCTGCGCGCG CCTGAAGAAG 551CCTTTGGCGA AGGCACGTCA CGTATGAAAA TTTGGACGGC ATCGGCTGAT 601GTAAATACTC CTTTTCAAAT CGGTAAACAG CTATTTGCCT ATGACACATC 651CGTTCATGCA CAATGGAACA AAACCCCGCT AACATCGCAA GACAAACTGG 701CTATCGGCGG ACACCACACC GTACGTGGCT TCGACGGTGA AATGAGTTTG 751CCTGCCGAGC GGGGATGGTA TTGGCGCAAC GATTTGAGCT GGCAATTTAA 801ACCAGGCCAT CAGCTTTATC TTGGGGCTGA TGTAGGACAT GTTTCAGGAC 851AATCCGCCAA ATGGTTATCG GGCCAAACTC TAGCCGGCAC AGCAATTGGG 901ATACGCGGGC AGATAAAGCT TGGCGGCAAC CTGCATTACG ATATATTTAC 951CGGCCGTGCA TTGAAAAAGC CCGAATATTT TCAGACGAAG AAATGGGTAA 1001 CGGGGTTTCA GGTGGGTTAT TCGTTTTGA

This encodes a protein having amino acid sequence <SEQ ID 608>:

  1 MDNSGSEATG KYQGNITFSA DNPFGLSDMF YVNYGRSIGG TPDEENFDGH  51RKEGGSNNYA VHYSAPFGKW TWAFNHNGYR YHQAVSGLSE VYDYNGKSYN 101TDFGFNRLLY RDAKRKTYLS VKLWTRETKS YIDDAELTVQ RRKTTGWLAE 151LSHKGYIGRS TADFKLKYKH GTGMKDALRA PEEAFGEGTS RMKIWTASAD 201VNTPFQIGKQ LFAYDTSVHA QWNKTPLTSQ DKLAIGGHHT VRGFDGEMSL 251PAERGWYWRN DLSWQFKPGH QLYLGADVGH VSGQSAKWLS GQTLAGTAIG 301IRGQIKLGGN LHYDIFTGRA LKKPEYFQTK KWVTGFQVGY SF*

The underlined sequence (aromatic-Xaa-aromatic amino acid motif) isusually found at the C-terminal end of outer membrane proteins.

ORF142ng and ORF142-1 show 95.6% identity over 342aa overlap:

In addition, ORF142ng is homologous to the HecB protein of E.chrysanthemi:

gi|1772622 (L39897) HecB [Erwinia chrysanthemi] Length = 558 Score =119 bits (295), Expect = 3e−26 Identities = 88/346 (25%), Positives =151/346 (43%), Gaps = 22/346 (6%) Query:   2DNSGSEATGKYQGNITFSADNPFGLSDMFYVNYGRSIGGTPDEENFDGHRKEGGSNNYAV  61DNSG ++TG+ Q N + + DN FGL+D ++++ G S   +    + D    + G Sbjct: 230DNSGQKSTGEEQLNGSLALDNVFGLADQWFISAGHS---SRFATSHDAESLQAG------ 280 Query: 62 HYSAPFGKWTWAFNHNGYRYHQAVSGLSEVYDYNGKSYNTDFGFNRLLYRDAKRKTYLSV 121 +S P+G W   +N++  RY          +   G S    F  +R+++RD   KT ++ Sbjct: 281-FSMPYGYWNLGYNYSQSRYRNTFINRDFPWHSTGDSDTHRFSLSRVVFRDGTMKTAIAG 339 Query:122 KLWTRETKSYIDDAELTVQRRKTTGWLAELSHKGYIGRSTADFKLKYKHGTGMKDALRAP 181    R   +Y++ + L    RK +     ++H   +    A F   Y  G     + Sbjct: 340TFSQRTGNNYLNGSLLPSSSRKLSSVSLGVNHSQKLWGGLATFNPTYNRGVRWLGSETDT 399 Query:182 EEAFGEGTSRMKIWTASADVNTPFQIGKQLFAYDTSVHAQWNKTPLTSQDKLAIGGHHTV 241 +++ E  +    WT SA    P         Y  S++ Q++   L   ++L +GG  ++ Sbjct: 400DKSADEPRAEFNKWTLSASYYHPV---TDSITYLGSLYGQYSARALYGSEQLTLGGESSI 456 Query:242 RGFDGEMSLPAERGWYWRNDLSWQFKP----GHQLYLGA-DVGHVSGQSAKWLSGQTLAG 296RGF  E      RG YWRN+L+WQ       G+  ++ A D GH+        +  +L G Sbjct: 457RGF-REQYTSGNRGAYWRNELNWQAWQLPVLGNVTFMAAVDGGHLYNHKQDNSTAASLWG 515 Query:297 TAIGIRGQIKLGGNLHYDIFTGRALKKPEYFQTKKWVTGFQVGYSF 342  A+G+    +   L   +  G  +  P + Q    V G++VG SF Sbjct: 516GAVGMTVASRW---LSQQVTVGWPISYPAWLQPDTMVVGYRVGLSF 558

On the basis of this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 73

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 609>:

  1 ATGCGGACGA AATGGTCAGC AGTGAGAAGC TGCTTACTTG GgCGGACACC  51GCCGACATCG ATACCGCTTT GAACCTGTTG TACCGTTTGC AAAAACTCGA 101ATTCCTCTAT GGCGATGAAA ACGGTCATTC AGACGGCATC AATTTGwCGG 151ACGAGCAATT GCCGTTGCTG ATGGAACAAT TGTCCGGCAG CGGTAAGGCG 201TTATTGGTCG ATCGGAACGG TCTGTATCTT GCCAACGCCA ATTTCCATCA 251TGAGGCGGCG GAAGAGTTGG GGTTGTTGGC GGCAGAAGTC GCACAGATGG 301AAAAGAAATA CCGGCTGCTG ATTAAGAACA AC..

This corresponds to the amino acid sequence <SEQ ID 610; ORF143>:

  1 MRTKWSAVRS CTWADTADID TALNLLYRLQ KLEFLYGDEN GHSDGINLXD  51EQLPLLMEQL SGSGKALLVD RNGLYLANAN FHHEAAEELG LLAAEVAQME 101 KKYRLLIKNN ..

Further work revealed the complete nucleotide sequence <SEQ ID 611>:

1 ATGGAATCAA CACTTTCACT ACAAGCAAAT TTATATCCCC GCCTGACTCC 51 TGCCGGTGCATTTTATGCCG TATCCAGCGA TGCCCCCAGT GCCGGTAAAA 101 CTTTGTTGCA CAGCCTGTTGAAAGCAGATG CGGACGAAAT GGTCAGCAGT 151 GAGAAGCTGC TTACTTGGGC GGACACCGCCGACATCGATA CCGCTTTGAA 201 CCTGTTGTAC CGTTTGCAAA AACTCGAATT CCTCTATGGCGATGAAAACG 251 GTCATTCAGA CGGCATCAAT TTGTCGGACG AGCAATTGCC GTTGCTGATG301 GAACAATTGT CCGGCAGCGG TAAGGCGTTA TTGGTCGATC GGAACGGTCT 351GTATCTTGCC AACGCCAATT TCCATCATGA GGCGGCGGAA GAGTTGGGGT 401 TGTTGGCGGCAGAAGTCGCA CAGATGGAAA AGAAATACCG GCTGCTGATT 451 AAGAACAACC TGTATATCAACAATAACGCT TGGGGCGTTT GCGATCCTTC 501 CGGTCAGAGC GAATTGACAT TTTTCCCATTGTATATCGGT TCAACCAAAT 551 TTATTTTGGT TATCGGCGGC ATTCCCGATT TGGGCAAAGAGGCATTTGTT 601 ACTTTGGTAA GGATTTTATA CCGCCGTTAC AGCAACCGCG TGTAA

This corresponds to the amino acid sequence <SEQ ID 612; ORF143-1>:

1 MESTLSLQAN LYPRLTPAGA FYAVSSDAPS AGKTLLHSLL KADADEMVSS 51 EKLLTWADTADIDTALNLLY RLQKLEFLYG DENGHSDGIN LSDEQLPLLM 101 EQLSGSGKAL LVDRNGLYLANANFHHEAAE ELGLLAAEVA QMEKKYRLLI 151 KNNLYINNNA WGVCDPSGQSELTFFPLYIG STKFILVIGG IPDLGKEAFV 201 TLVRILYRRY SNRV*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF143 shows 92.4% identity over a 105aa overlap with an ORF (ORF143a)from strain A of N. meningitidis:

The complete length ORF143a nucleotide sequence <SEQ ID 613> is:

1 ATGGAATCAA CANTTTCACT ACAAGCAAAT TTATATCNCC GCCTGACTCC 51 TGCCGGTGCATTTTATGCCG TATCCAGCGA TGNCCCCAGT GCCGGTAAAA 101 CTTTGTTGCA CAGCCTGTTGAAAGCGGATG CGGACGAAAT GGTNAGCAGT 151 GAGAAGCTGC TTACCTGGGC GGANACCGCCGACATCGATA CCGCTTTGAA 201 CCTGTTGTAC CGTTTGCAAA AACTCGAATT CCTCTATGGCGATGAAAACG 251 GTCATTCAGA CGGCATCAAT TTGTCGGACG AGCAATTGCC GTTGCTGATG301 GAACAATTGT CCGGCAGCGG TAAGGCGTTA TTGGTCGATC GGAACGGTCT 351GTATCTTGCC AACGCCAATT TCCATCATGA GGCGGCGGAA GAGTTGGGGT 401 TGTTGGCGGCAGAAGTCGCA CAGATGGAAA AGAAATACCG GCTGCNNATT 451 AAGAACAACC TGTATATCAACAATAACGCT TGGGGCGTTT GCGATCCTTC 501 CGGTCAGAGC GAATTGACAT TTTTCCCATTGTATATCGGT TCAACCAAAT 551 TTATTTTGGT TATCGGCGGC ATTCCCGATT TGGGCAAAGAGGCATTTGTT 601 ACTTTGGTAA GGATNTTATA CCNCCNGTTA CAGCAACCGC GTGTAAAACT651 TGGGAGAGAG GANGGGTTAT GCAGCAATTA TTGA

This encodes a protein having amino acid sequence <SEQ ID 614>:

1 MESTXSLQAN LYXRLTPAGA FYAVSSDXPS AGKTLLHSLL KADADEMVSS 51 EKLLTWAXTADIDTALNLLY RLQKLEFLYG DENGHSDGIN LSDEQLPLLM 101 EQLSGSGKAL LVDRNGLYLANANFHHEAAE ELGLLAAEVA QMEKKYRLXI 151 KNNLYINNNA WGVCDPSGQSELTFFPLYIG STKFILVIGG IPDLGKEAFV 201 TLVRXLYXXL QQPRVKLGRE XGLCSNY*

ORF143a and ORF143-1 show 97.1% identity in 207 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF143 shows 95.5% identity over a 110aa overlap with a predicted ORF(ORF143ng) from N. gonorrhoeae:

An ORF143ng nucleotide sequence <SEQ ID 615> was predicted to encode aprotein having amino acid sequence <SEQ ID 616>:

1 MRTKWSAVRS CSRADTADID TALNLLYRLQ KLEFLYGDEN GHSDGINLSD 51 EQLPLLMEQLSGSGKALLVD RNGLYLANAN FHHESAEELG LLAAEVAQME 101 KKYRLLIRNN LYINNNAWGVCDPSGQSELT FFPLYIGSTK FILVIAGIPD 151 LSKGGICYFG KDFIPPLQQP RVKLGTGGIMRQLLISILED LNNTSTDIIA 201 SAVISTDGLP MATMLPSHLN SDRVGAISAT LLALGSRSVQELACGELEQV 251 MIKGKSGYIL LSQAGKDAVL VLVAKETGRL GLILLDAKRA ARHIAEAI*

Further work revealed the following gonococcal DNA sequence <SEQ ID617>:

1 ATGGAATCAA CACTTTCACT ACAAGCGAAT TTATATCCCT GCCTGACTCC 51 TGCCGGTGCATTTTATGCCG TATCCAGCGA TGCCCCCAGT GCCGGTAAAA 101 CTTTGTTGCG CAGCCTGTTGAAAGCGGATG CGGACGAAGT GGTCAGCAGT 151 GAGAAGCTGC TCGCGGCGGA CACCGCCGACATCGATACCG CTTTGAACCT 201 GTTGTACCGT TTGCAAAAAC TCGAATTCCT CTATGGCGATGAAAACGGTC 251 ATTCAGACGG CATCAATTTG TCGGACGAGC AATTGCCGTT GCTGATGGAA301 CAATTGTCCG GCAGCGGTAA GGCATTATTG GTCGATCGGA ACGGTCTGTA 351TCTTGCCAAC GCCAATTTCC ATCATGAGTC GGCGGAAGAG TTGGGGTTGT 401 TGGCGGCAGAAGTCGCACAG ATGGAAAAGA AATACCGGCT GCTGATTAGG 451 AACAACCTGT ATATCAACAATAACGCTTGG GGCGTTTGCG ATCCTTCCGG 501 TCAGAGCGAA TTGACATTTT TCCCATTGTATATCGGTTCA ACCAAATTTA 551 TTTTGGTTAT CGCCGGCATT CCCGATTTGA GCAAAGAGGCATTTGTTACT 601 TTGGTAAGGA TTTTATACCG CCGTTACAGC AACCGCGTGT AA

This corresponds to the amino acid sequence <SEQ ID 618; ORF143ng-1>:

1 MESTLSLQAN LYPCLTPAGA FYAVSSDAPS AGKTLLRSLL KADADEVVSS 51 EKLLAADTADIDTALNLLYR LQKLEFLYGD ENGHSDGINL SDEQLPLLME 101 QLSGSGKALL VDRNGLYLANANFHHESAEE LGLLAAEVAQ MEKKYRLLIR 151 NNLYINNNAW GVCDPSGQSELTFFPLYIGS TKFILVIAGI PDLSKEAFVT 201 LVRILYRRYS NRV*

ORF143ng-1 and ORF143-1 show 95.8% identity in 214 aa overlap:

Based on the presence of the putative transmembrane domains in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 74

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 619>:

1 ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA AAATCTGTGC 51 GTTTGCATGGTTCGTCGTCC GCCGCTTTGA TGAAGAACGC GTACCGCAGr 101 CGGCGGCAAG CATGACGTTTACGACGCTGC TGGCACTCGT CCCCGTGCTG 151 ACCGTGATGG TGGCGGTCGC TTCGATTTTCCCCGTGTTCG ACCGCTGGTC 201 GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCGCA.GGCGCGG 251 ACATGGTGTT CGACTATATC AATGCGTTCC GCGAGCAGGC GAACCGGCTG301 ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCTGA TGCTGATTCG 351GACGATAGAC AATACGTTCA ACCGCATCTG GaCGGGTCAA wTyCCAGCGT 401 CCGTGGATG..

This corresponds to the amino acid sequence <SEQ ID 620; ORF144>:

1 MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQXAASMTF TTLLALVPVL 51 TVMVAVASIFPVFDRWSDSF VSFVNQTIVP XGADMVFDYI NAFREQANRL 101 TAIGSVMLVV TSLMLIRTIDNTFNRIWRVX XQRPWM...

Further work revealed the complete nucleotide sequence <SEQ ID 621>:

1 ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA AAATCTGTGC 51 GTTTGCATGGTTCGTCGTCC GCCGCTTTGA TGAAGAACGC GTACCGCAGG 101 CGGCGGCAAG CATGACGTTTACGACGCTGC TGGCACTCGT CCCCGTGCTG 151 ACCGTGATGG TGGCGGTCGC TTCGATTTTCCCCGTGTTCG ACCGCTGGTC 201 GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCGCAGGGCGCGG 251 ACATGGTGTT CGACTATATC AATGCGTTCC GCGAGCAGGC GAACCGGCTG301 ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCTGA TGCTGATTCG 351GACGATAGAC AATACGTTCA ACCGCATCTG GCGGGTCAAT TCCCAGCGTC 401 CGTGGATGATGCAGTTTCTC GTCTATTGGG CTTTACTGAC GTTCGGGCCG 451 CTGTCTTTGG GCGTGGGCATTTCCTTTATG GTCGGCTCGG TACAGGATGC 501 CGCGCTTGCC TCAGGTGCGC CGCAGTGGTCGGGCGCGTTG CGAACGGCGG 551 CGACGCTGAC CTTCATGACG CTTTTGCTGT GGGGGCTGTACCGCTTCGTG 601 CCAAACCGCT TCGTTCCCGC GCGGCAGGCG TTTGTCGGGG CTTTGGCAAC651 AGCGTTTTGT CTGGAAACCG CGCGCTCCCT CTTCACTTGG TATATGGGCA 701ATTTCGACGG CTACCGCTCG ATTTACGGCG CGTTTGCCGC CGTGCCGTTT 751 TTTCTGTTGTGGCTGAACCT GTTGTGGACG CTGGTCTTGG GCGGCGCGGT 801 GCTGACTTCT TCACTCTCCTACTGGCAGGG AGAAGCGTTC CGCAGGGGCT 851 TCGACTCGCG CGGACGGTTT GACGACGTGTTGAAAATCCT GCTGCTTCTG 901 GATGCGGCGC AAAAAGAAGG CAAAGCCTTG CCTGTTCAGGAGTTCAGACG 951 GCATATCAAT ATGGGCTACG ACGAGTTGGG CGAGCTTTTG GAAAAGCTGG1001 CGCGGCACGG CTACATCTAT TCCGGCAGAC AGGGTTGGGT GTTGAAAACG 1051GGGGCGGATT CGATTGAGTT GAACGAACTC TTCAAGCTCT TCGTTTACCG 1101 TCCGTTGCCTGTGGAAAGGG ATCATGTGAA CCAAGCTGTC GATGCGGTAA 1151 TGACACCGTG TTTGCAGACTTTGAACATGA CGCTGGCAGA GTTTGACGCT 1201 CAGGCGAAAA AACGGCAGTA G

This corresponds to, the amino acid sequence <SEQ ID 622; ORF144-1>:

1 MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQAAASMTF TTLLALVPVL 51 TVMVAVASIFPVFDRWSDSF VSFVNQTIVP QGADMVFDYI NAFREQANRL 101 TAIGSVMLVV TSLMLIRTIDNTFNRIWRVN SQRPWMMQFL VYWALLTFGP 151 LSLGVGISFM VGSVQDAALA SGAPQWSGALRTAATLTFMT LLLWGLYRFV 201 PNRFVPARQA FVGALATAFC LETARSLFTW YMGNFDGYRSIYGAFAAVPF 251 FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF DDVLKILLLL301 DAAQKEGKAL PVQEFRRHIN MGYDELGELL EKLARHGYIY SGRQGWVLKT 351GADSIELNEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT LNMTLAEFDA 401 QAKKRQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF144 shows 96.3% identity over a 136aa overlap with an ORF (ORF144a)from strain A of N. meningitidis:

The complete length ORF144a nucleotide sequence <SEQ ID 623> is:

1 ATGACCTTTT TACAACGTTT GCAAGGTTTG GCAGACAATA AAATCTGTGC 51 GTTTGCATGGTTCGTCGTCC GCCGCTTTGA TGAAGAACGC GTACCGCAGG 101 CGGCGGCAAG CATGACGTTTACGACACTGC TGGCACTCGT CCCCGTGCTG 151 ACCGTGATGG TGGCGGTCGC TTCGATTTTCCCCGTGTTCG ACCGNTGGTC 201 GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCGCAGGGCGCGG 251 ACATGGTNTT CGACTATATC AATGCGTTCC GCGAGCAGGC GAACCGGCTG301 ACGGCAATCG GCAGCGTGAT GCTGGTCGTT ACCTCGCNGA TGCTGATTCG 351GACGATAGAC AATACGTTCA ACCGCATCTG GCGGGTCAAT TCCCAGCGTC 401 CGTGGATGATGCAGTTTCTC GTCTATTGGG CTTTACTGAC GTTCGGGCCG 451 CTGTCTTTGG GCGTGGGCATTTCCTTTATN GTCGGCTCGG TACAGGATGC 501 CGCGCTTGCC TCAGGTGCGC CGCAGTGGTCGGGCGCGTTG CGAACGGCGG 551 CGACGCTGAN CTTCATGACG CTTTTGCTGT GGGGGCTGTACCGCTNCGTG 601 CCAAACCGCT TCGTTCCCGC GCGGCANGCG TTTGTCGGGG CTTTGGCAAC651 AGCGTTCTGT CTGGAAACCG CGCGTTCCCT CTTTACTTGG TATATGGGCA 701ATTTCGACGG CTACCGCTCG ATTTACGGNG CGTTTGCCGC CGTGCCGTTT 751 TTTCTGTTGTGGCTGAACCT GTTGTGGACG CTGGTCTTGG GCGGCGCGGT 801 GCTGACTTCT TCACTCTCCTACTGGCAGGG AGAAGCGTTC CGCAGGGNCT 851 TCGACTCGCG CGGACGGTTT GACGACGTGTTGAAAATCCT GCTGCTTCTG 901 GATGCGGCGC AAAAAGAAGG CNAAGCCTTG CCTGTTCAGGAGTTCAGACG 951 GCATATCAAT ATGGGCTACG ACGAGTTGGG CGAGCTTTTG GAAAAGCTGG1001 CGCGGCACGG CTACATCTAT TCCGGCAGAC AGGGTTGGGT GTTGAAAACG 1051GGGGCGGATT CGATTGAGTT GAACGAACTC TTCAAGCTCT TCGTTTACCG 1101 TCCGTTGCCTGTGGAAAGGG ATCATGTGAA CCAAGCTGTC GATGCGGTAA 1151 TGATGCCGTG TTTGCAGACTTTGAACATGA CGCTGGCAGA GTTTGACGCT 1201 CAGGCGAAAA AACAGCAGCA ATCTTGA

This encodes a protein having amino acid sequence <SEQ ID 624>:

1 MTFLQRLQGL ADNKICAFAW FVVRRFDEER VPQAAASMTF TTLLALVPVL 51 TVMVAVASIFPVFDRWSDSF VSFVNQTIVP QGADMVFDYI NAFREQANRL 101 TAIGSVMLVV TSXMLIRTIDNTFNRIWRVN SQRPWMMQFL VYWALLTFGP 151 LSLGVGISFX VGSVQDAALA SGAPQWSGALRTAATLXFMT LLLWGLYRXV 201 PNRFVPARXA FVGALATAFC LETARSLFTW YMGNFDGYRSIYGAFAAVPF 251 FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRXFDSRGRF DDVLKILLLL301 DAAQKEGXAL PVQEFRRHIN MGYDELGELL EKLARHGYIY SGRQGWVLKT 351GADSIELNEL FKLFVYRPLP VERDHVNQAV DAVMMPCLQT LNMTLAEFDA 401 QAKKQQQS*

ORF144a and ORF144-1 show 97.8% identity in 406 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF144 shows 91.2% identity over a 136aa overlap with a predicted ORF(ORF144ng) from N. gonorrhoeae:

The complete length ORF144ng nucleotide sequence <SEQ ID 625> ispredicted to encode a protein having amino acid sequence <SEQ ID 626>:

1 MTFLQCWQGS ADNKICAFAW FVIRRFSEER VPQAAASMTF TTLLALVPVL 51 TVMVAVASIFPVFDRWSDSF VSFVNQTIVP QGADMVFDYI DAFRDQANRL 101 TAIGSVMLVV TSLMLIRTIDNAFNRIWRVN TQRPWMMQFL VYWALLTFGP 151 LSLGVGISFM VGSVQDSVLS SGAQQWADALKTAARLAFMT LLLWGLYRFV 201 PNRFVPARQA FVGALITAFC LETARFLFTW YMGNFDGYRSIYGAFAAVPF 251 FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF DDVLKILLLL301 DAAQKEGRTL SVQEFRRHIN MGYDELGELL EKLARYGYIY SGRQGWVLKT 351GADSIELSEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT LNMTLAEFDA 401 QAKKQQQS*

Further work revealed the following gonococcal DNA sequence <SEQ ID627>:

1 ATGACCTTTT TACAACGTTG GCAAGGTTTG GCGGACAATA AAATCTGTGC 51 ATTTGCATGGTTCGTCATCC GCCGTTTCAG TGAAGAGCGC GTACCGCAGG 101 CAGCGGCGAG CATGACGTTTACGACACTGC TGGCACTCGT CCCCGTACTG 151 ACCGTAATGG TCGCGGTCGC TTCGATTTTCCCCGTGTTCG ACCGCTGGTC 201 GGATTCGTTC GTCTCCTTCG TCAACCAAAC CATTGTGCCGCAGGGCGCGG 251 ATATGGTGTT CGACTATATC GACGCATTCC GCGATCAGGC AAACCGGCTG301 ACCGCCATCG GCAGCGTGAT GCTGGTCGTA ACCTCGCTGA TGCTGATTCG 351GACGATAGAC AATGCGTTCA ACCGCATCTG GCGGGTTAAC ACGCAACGCC 401 CCTGGATGATGCAGTTCCTC GTTTATTGGG CGTTGCTGAC TTTCGGGCCT 451 TTGTCTTTGG GTGTGGGCATTTCCTTTATG GTCGGGTCGG TTCAAGACTC 501 CGTACTCTCC TCCGGAGCGC AACAATGGGCGGACGCGTTG AAGACGGCGG 551 CAAGGCTGGC TTTCATGACG CTTTTGCTGT GGGGGCTGTACCGCTTCGTG 601 CCCAACCGCT TCGTGCCCGC CCGGCAGGCG TTTGTCGGAG CTTTGATTAC651 GGCATTCTGC CTGGAGACGG CACGTTTCCT GTTCACCTGG TATATGGGCA 701ATTTCGACGG CTACCGCTCG ATTTACGGCG CATTTGCCGC CGTGCCGTTT 751 TTCCTGCTGTGGTTAAACCT GCTGTGGACG CTGGTCTTGG GCGGGGCGGT 801 GCTGACTTCG TCGCTGTCTTATTGGCAGGG CGAGGCCTTC CGCAGGGGAT 851 TCGACTCGCG CGGACGGTTT GACGACGTGTTGAAAATCCT GCTGCTTCTG 901 GATGCGGCGC AAAAAGAAGG CCGAACCCTG TCCGTTCAGGAGTTCAGACG 951 GCATATCAAT ATGGGTTACG ATGAATTGGG CGAGCTTTTG GAAAAGCTGG1001 CGCGGTACGG CTATATCTAT TCCGGCAGAC AGGGCTGGGT TTTGAAAACG 1051GGGGCGGATT CGATTGAGTT GAGCGAACTC TTCAAGCTCT TCGTGTACCG 1101 CCCGTTGCctgtggaAAGGG ATCATGTGAA CCAAGCTGtc gaTGCGGTAA 1151 TGAcgccgtG TTTGCAGACTTTGAACATGA CGCTGGCGGA GTTTGACGCT 1201 CAGgcgAAAA AACAGCAGCA GTCTTGA

This encodes a variant of ORF144ng, having the amino acid sequence <SEQID 628; ORF144ng-1>:

1 MTFLQCWQGL ADNKICAFAW FVIRRFSEER VPQAAASMTF TTLLALVPVL 51 TVMVAVASIFPVFDRWSDSF VSFVNQTIVP QGADMVFDYI DAFRDQANRL 101 TAIGSVMLVV TSLMLIRTIDNAFNRIWRVN TQRPWMMQFL VYWALLTFGP 151 LSLGVGISFM VGSVQDSVLS SGAQQWADALKTAARLAFMT LLLWGLYRFV 201 PNRFVPARQA FVGALITAFC LETARFLFTW YMGNFDGYRSIYGAFAAVPF 251 FLLWLNLLWT LVLGGAVLTS SLSYWQGEAF RRGFDSRGRF DDVLKILLLL301 DAAQKEGRTL SVQEFRRHIN MGYDELGELL EKLARYGYIY SGRQGWVLKT 351GADSIELSEL FKLFVYRPLP VERDHVNQAV DAVMTPCLQT LNMTLAEFDA 401 QAKKQQQS*

ORF144ng-1 and ORF144-1 show 94.1% identity in 406 aa overlap:

On this basis of this analysis, including the identification of severalputative transmembrane domains in the gonococcal protein, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 75

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 629>:

1 ..AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACC CCGAACTGGA 51  AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC CTCTGGCTCA 101  GCACCGATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT GCTGCAACGC 151  ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC TGCGCCAAAG 201  CCTGCTTGAA ACACGGGAAC ACGGCTGA

This corresponds to the amino acid sequence <SEQ ID 630; ORF146>:

1 ..RHARRIRIDT AINPELEALA EHLHYQWQGF LWLSTDMRQE ISALVILLQR 51  TRRKWLDAHE RQHLRQSLLE TREHG*

Further work revealed the complete nucleotide sequence <SEQ ID 631>:

1 ATGAACACCT CGCAACGCAA CCGCCTCGTC AGCCGCTGGC TCAACTCCTA 51 CGAACGCTACCGCTACCGCC GCCTCATCCA CGCCGTCCGG CTCGGCGGGG 101 CCGTCCTGTT CGCCACCGCCTCCGCCCGGC TGCTCCACCT CCAACACGGC 151 GAGTGGATAG GGATGACCGT CTTCGTCGTCCTCGGCATGC TCCAGTTTCA 201 AGGGGCGATT TACTCCAAGG CGGTGGAACG TATGCTCGGCACGGTCATCG 251 GGCTGGGCGC GGGTTTGGGC GTTTTATGGC TGAACCAGCA TTATTTCCAC301 GGCAACCTCC TCTTCTACCT CACCGTCGGC ACGGCAAGCG CACTGGCCGG 351CTGGGCGGCG GTCGGCAAAA ACGGCTACGT CCCTATGCTG GCAGGGCTGA 401 CGATGTGTATGCTCATCGGC GACAACGGCA GCGAATGGCT CGACAGCGGA 451 CTCATGCGCG CCATGAACGTCCTCATCGGC GCGGCCATCG CCATCGCCGC 501 CGCCAAACTG CTGCCGCTGA AATCCACACTGATGTGGCGT TTCATGCTTG 551 CCGACAACCT GGCCGACTGC AGCAAAATGA TTGCCGAAATCAGCAACGGC 601 AGGCGCATGA CCCGCGAACG CCTCGAGGAG AACATGGCGA AAATGCGCCA651 AATCAACGCA CGCATGGTCA AAAGCCGCAG CCATCTCGCC GCCACATCGG 701GCGAAAGCCG CATCAGCCCC GCCATGATGG AAGCCATGCA GCACGCCCAC 751 CGTAAAATCGTCAACACCAC CGAGCTGCTC CTGACCACCG CCGCCAAGCT 801 GCAATCTCCC AAACTCAACGGCAGCGAAAT CCGGCTGCTT GACCGCCACT 851 TCACACTGCT CCAAACCGAC CTGCAACAAACCGTCGCCCT TATCAACGGC 901 AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACCCCGAACTGGA 951 AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC CTCTGGCTCA1001 GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT GCTGCAACGC 1051ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC TGCGCCAAAG 1101 CCTGCTTGAAACACGGGAAC ACGGCTGA

This corresponds to the amino acid sequence <SEQ ID 632; ORF146-1>:

1 MNTSQRNRLV SRWLNSYERY RYRRLIHAVR LGGAVLFATA SARLLHLQHG 51EWIGMTVFVV LGMLQFQGAI YSKAVERMLG TVIGLGAGLG VLWLNQHYFH 101 GNLLFYLTVGTASALAGWAA VGKNGYVPML AGLTMCMLIG DNGSEWLDSG 151LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLADC SKMIAEISNG 201 RRMTRERLEENMAKMRQINA RMVKSRSHLA ATSGESRISP AMMEAMQHAH 251 RKIVNTTELL LTTAAKLQSPKLNGSEIRLL DRHFTLLQTD LQQTVALING 301 RHARRIRIDT AINPELEALA EHLHYQWQGFLWLSTNMRQE ISALVILLQR 351 TRRKWLDAHE RQHLRQSLLE TREHG*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF146 shows 98.6% identity over a 74aa overlap with an ORF (ORF146a)from strain A of N. meningitidis:

The complete length ORF146a nucleotide sequence <SEQ ID 633> is:

1 ATGAACACCT CGCAACGCAA CCGCCTCGTC AGCCGCTGGC TCAACTCCTA 51 CGAACGCTACCGCTACCGCC GCCTCATCCA CGCCGTCCGG CTCGGCGGGG 101 CCGTCCTGTT CGCCACCGCCTCCGCCCGGC TGCTCCACCT CCAACACGGC 151 GAGTGGATAG GGATGACCGT CTTCGTCGTCCTCGGCATGC TCCAGTTTCA 201 AGGGGCGATT TACTCCAAGG CGGTGGAACG TATGCTCGGCACGGTCATCG 251 GGCTGGGCGC GGGTTTGGGC GTTTTATGGC TGAACCAGCA TTATTTCCAC301 GGCAACCTCC TCTTCTACCT CACCGTCGGC ACGGCAAGCG CACTGGCCGG 351CTGGGCGGCG GTCGGCAAAA ACGGCTACGT CCCTATGCTG GCGGGGCTGA 401 CGATGTGCATGCTCATCGGC GACAACGGCA GCGAATGGTT CGACAGCGGC 451 CTGATGCGCG CGATGAACGTCCTCATCGGC GCGGCCATCG CCATCGCCGC 501 CGCCAAACTG CTGCCGCTGA AATCCACACTGATGTGGCGT TTCATGCTTG 551 CCGACAACCT GACCGACTGC AGCAAAATGA TTGCCGAAATCAGCAACGGC 601 AGGCGCATGA CCCGCGAACG CCTCGAAGAG AACATGGCGA AAATGCGCCA651 AATCAACGCA CGCATGGTCA AAAGCCGCAG CCACCTCGCC GCCACATCGG 701GCGAAAGCCG CATCAGCCCC GCCATGATGG AAGCCATGCA GCACGCCCAC 751 CGTAAAATTGTCAACACCAC CGAGCTGCTC CTGACCACCG CCGCCAAGCT 801 GCAATCTCCC AAACTCAACGGCAGCGAAAT CCGGCTGCTT GACCGCCACT 851 TCACACTGCT CCAAACCGAC CTGCAACAAACCGTCGCCCT TATCAACGGC 901 AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACCCCGAACTGGA 951 AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC CTCTGGCTCA1001 GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT GCTGCAACGC 1051ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC TGCGCCAAAG 1101 CCTGCTTGAAACACGGGAAC ACAGTTGA

This encodes a protein having amino acid sequence <SEQ ID 634>:

1 MNTSQRNRLV SRWLNSYERY RYRRLIHAVR LGGAVLFATA SARLLHLQHG 51EWIGMTVFVV LGMLQFQGAI YSKAVERMLG TVIGLGAGLG VLWLNQHYFH 101 GNLLFYLTVGTASALAGWAA VGKNGYVPML AGLTMCMLIG DNGSEWFDSG 151LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLTDC SKMIAEISNG 201 RRMTRERLEENMAKMRQINA RMVKSRSHLA ATSGESRISP AMMEAMQHAH 251 RKIVNTTELL LTTAAKLQSPKLNGSEIRLL DRHFTLLQTD LQQTVALING 301 RHARRIRIDT AINPELEALA EHLHYQWQGFLWLSTNMRQE ISALVILLQR 351 TRRKWLDAHE RQHLRQSLLE TREHS*

ORF146a and ORF146-1 show 99.5% identity in 374 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF146 shows 97.3% identity over a 75aa overlap with a predicted ORF(ORF146ng) from N. gonorrhoeae:

An ORF146ng nucleotide sequence <SEQ ID 635> was predicted to encode aprotein having amino, acid sequence <SEQ ID 636>:

1 MSGVRFPSPA PIPSTDPPSG SLCFFTFPLQ TASDMNSSQR KRLSGRWLNS 51 YERYRHRRLIHAVRLGGTVL FATALARLLH LQHGEWIGMT VFVVLGMLQF 101 QGAIYSNAVERMLGTVIGLG AGLGVLWLNQ HYFHGNLLFY LTIGTASALA 151 GWAAVGKNGY VPMLAGLTMCMLIGDNGSEW LDSGLMRAMN VLIGAAIAIA 201 AAKLLPLKST LMWRFMLADN LADCSKMIAEISNGRRMTRE RLEQNMVKMR 251 QINARMVKSR SHLAATSGES RISPSMMEAM QHAHRKIVNTTELLLTTAAK 301 LQSPKLNGSE IRLLDRHFTL LQTDLQQTAA LINGRHARRI RIDTAINPEL351 EALAEHLHYQ WQGFLWLSTN MRQEISALVI PLQRTRRKWL DAHERQHLRQ 401SLLETREHG*

Further work revealed the following gonococcal DNA sequence <SEQ ID637>:

1 ATGAACTCCT CGCAACGCAA ACGCCTTTCC GgccGCTGGC TCAACTCCTA 51 CGAACGCTaccGCCaccGCC GCCTCATACA TGCCGTGCGG CTCGGCggaa 101 ccgtCCTGTT CGCCACCGCACTCGCCCGgc tACTCCACCT CCAacacggc 151 gAATGGATAG GGAtgaCCGT CTTCGTCGTCCTCGGCATGC TCCAGTTCCA 201 AGGCgcgatt tActccaacg cggtgGAacg taTGctcggtacggtcatcg 251 ggctgGGCGC GGGTTTGGgc gTTTTATGGC TGAACCAGCA TTAtttccac301 ggcaacCTcc tcttctacct gaccatcggc acggcaagcg cactggccgg 351ctGGGCGGCG GTCGGCAAAA acggctacgt ccctatgctg GCGGGGctgA 401 CGATGTGCATgctcatcggc gACAACGGCA GCGAATGGCT CGACAGCGGC 451 CTGATGCGCG CGATGAACGTCCTCATCGGC GCCGCCATCG CCATTGCCGC 501 CGCCAAACTG CTGCCGCTGA AATCCACACTGATGTGGCGT TTCATGCTTG 551 CCGACAACCT GGCCGACTGC AGCAAAATGA TTGCCGAAATCAGCAACGGC 601 AGGCGTATGA CGCGCGAACG TTTGGAGCAG AATATGGTCA AAATGCGCCA651 AATCAACGCA CGCATGGTCA AAAGCCGCAG CCACCTCGCC GCCACATCGG 701GCGAAAGCCG CATCAGCCCC TCCATGATGG AAGCCATGCA GCACGCCCAC 751 CGCAAAATCGTCAACACCAC CGAGCTGCTC CTGACCACCG CCGCCAAGCT 801 GCAATCTCCC AAACTCAACGGCAGCGAAAT CCGGCTGCTC GACCGCCACT 851 TCACACTGCT CCAAACCGAC CTGCAACAAACCGCCGCCCT CATCAACGGC 901 AGACACGCCC GCCGCATCCG CATCGACACC GCCATCAACCCCGAACTGGA 951 AGCCCTCGCC GAACACCTCC ACTACCAATG GCAGGGCTTC CTCTGGCTCA1001 GCACCAATAT GCGTCAGGAA ATTTCCGCCC TCGTCATCCT GCTGCAACGC 1051ACCCGCCGCA AATGGCTGGA TGCCCACGAA CGCCAACACC TGCGCCAAAG 1101 CCTGCTTGAAACACGGGAAC ACGGCTGA

This corresponds to the amino acid sequence <SEQ ID 638; ORF146ng-1>:

1 MNSSQRKRLS GRWLNSYERY RHRRLIHAVR LGGTVLFATA LARLLHLQHG 51EWIGMTVFVV LGMLQFQGAI YSNAVERMLG TVIGLGAGLG VLWLNQHYFH 101 GNLLFYLTIGTASALAGWAA VGKNGYVPML AGLTMCMLIG DNGSEWLDSG 151LMRAMNVLIG AAIAIAAAKL LPLKSTLMWR FMLADNLADC SKMIAEISNG 201 RRMTRERLEQNMVKMRQINA RMVKSRSHLA ATSGESRISP SMMEAMQHAH 251 RKIVNTTELL LTTAAKLQSPKLNGSEIRLL DRHFTLLQTD LQQTAALING 301 RHARRIRIDT AINPELEALA EHLHYQWQGFLWLSTNMRQE ISALVILLQR 351 TRRKWLDAHE RQHLRQSLLE TREHG*

ORF146ng-1 and ORF146-1 show 96.5% identity in 375 aa overlap

Furthermore, ORF146ng-1 shows homology with a hypothetical E. coliprotein:

sp|P33011|YEEA_ECOLI HYPOTHETICAL 40.0 KD PROTEIN IN COBU-SBMCINTERGENIC REGION >gi|1736674|gnl|PID|d1016553 (D90838) ORF_ID: o348#20;similar to [SwissProt Accession Number P33011] [Escherichiacoli] >gi|1736682|gnl|PID|d1016560 (D90839) ORF_ID: o348#20; similar to[SwissProt Accession Number P33011] [Escherichia coli] >gi|1788318(AE000292) f352; 100% identical to fragment YEEA_ECOLI SW: P33011 buthas 203 additional C-terminal residues [Escherichia coli] Length = 352Score = 109 bits (271), Expect = 2e−23 Identities = 89/347 (25%),Positives = 150/347 (42%), Gaps = 21/347 (6%) Query: 20YRHRRLIHAVRLGGTVLFATALARLLHLQHGEWIGMTVFVVLGMLQFQGAIYSNAVERML 79 YRHR++H  R+    L    + RL  +    W  +T+ V++G + F G +   A ER+ Sbjct: 15YRHYRIVHGTRVALAFLLTFLIIRLFTIPESTWPLVTMVVIMGPISFWGNVVPRAFERIG 74 Query:80 GTVIGLGAGLGVLWLNQHYFHGNLLFYLTIGTASALAGWAAVGKNGYVPMLAGLTMCMLI 139GTV+G   GL  L L         L  +    A  L GW A+GK  Y  +L G+T+ +++ Sbjct: 75GTVLGSILGLIALQLE---LISLPLMLVWCAAAMFLCGWLALGKKPYQGLLIGVTLAIVV 131 Query:140 GDNGSEWLDSGLMRAMNVLIGXXXXXXXXKLLPLKSTLMWRFMLADNLADCSKMIAEISN 199G    E +D+ L R+ +V++G         + P ++ + WR  LA +L + +++     + Sbjct: 132GSPTGE-IDTALWRSGDVILGSLLAMLFTGIWPQRAFIHWRIQLAKSLTEYNRVYQSAFS 190 Query:200 GRRMTRERLEQNMVKMRQINARMVKSRSHLAATSGESRISPSMMEAMQHAHRKIVNXXXX 259   + R RLE ++ K+       VK R  +A  S E+RI  S+ E +Q  +R +V Sbjct: 191PNLLERPRLESHLQKLL---TDAVKMRGLIAPASKETRIPKSIYEGIQTINRNLVCMLEL 247 Query:260 XXXXXXXXQSPK---LNGSEIRLLDRHFXXXXXXXXXXAALINGRHARRIRIDTAINPEL 316        +      LN  ++R  D              AL  G           +N  + Sbjct: 248QINAYWATRPSHFVLLNAQKLR--DTQHMMQQILLSLVHALYEGNPQPVFANTEKLNDAV 305 Query:317 EALAEHL--HYQWQ-------GFLWLSTNMRQEISALVILLQRTRRK 354 E L+ L  H+  +       G++WL+     ++  L  L+ R  RK Sbjct: 306EELRQLLNNHHDLKVVETPIYGYVWLNMETAHQLELLSNLICRALRK 352

On the basis of this analysis, including the identification of severaltransmembrane domains in the gonococcal protein, it is predicted thatthe proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 76

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 639>

1 ..GCCGAAGACA CGCGCGTTAC CGCACAGCTT TTGAGCGCGT ACGGCATTCA 51  GGGCAAACTC GTCAGTGTGC GCGAACACAA CGAACGGCAG ATGGCGGACA 101  AGATTGTCGG CTATCTTTCA GACGGCATGG TTGTGGCACA GGTTTCCGAT 151  GCGGGTACGC CGGCCGTGTG CGACCCGGGC GCGAAACTCG CCCGCCGCGT 201  GCGTGAGGCC GGGTTTAAAG TCGTTCCCGT CGTGGGCGCA AC.GCGGTGA 251  TGGCGGCTTT GAGCGTGGCC GGTGTGGAAG GATCCGATTT TTATTTCAAC 301  GGTTTTGTAC CGCCGAAATC GGGAGAACGC AGGAAACTGT TTGCCAAATG 351  GGTGCGGGCG GCGTTTCCTA TCGTCATGTT TGAAACGCCG CACCGCATCG 401  GTGCAGCGCT TGCCGATATG GCGGAACTGT TCCCCGAACG CCGATTAATG 451  CTGGCGCGCG AAATTACGAA AACGTTTGAA ACGTTCTTAA GCGGCACGGT 501  TGGGGAAATT CAGACGGCAT TGTCTGCCGA CGGCGACCAA TCGCGCGGCG 551  AGATGGTGTT GGTGCTTTAT CCGGCGCAGG ATGAAAAACA CGAAGGCTTG 601  TCCGAGTCCG CGCAAAACAT CATGAAAATC CTCACAGCCG AGCTGCCGAC 651  CAAACAGGCG GCGGAGCTTG CTGCCAAAAT CACGGGCGAG GGAAAGAAAG 701  CTTTGTACGA T..

This corresponds to the amino acid sequence <SEQ ID 640; ORF147>:

1 ..AEDTRVTAQL LSAYGIQGKL VSVREHNERQ MADKIVGYLS DGMVVAQVSD 51  AGTPAVCDPG AKLARRVREA GFKVVPVVGA XAVMAALSVA GVEGSDFYFN 101  GFVPPKSGER RKLFAKWVRA AFPIVMFETP HRIGAALADM AELFPERRLM 151  LAREITKTFE TFLSGTVGEI QTALSADGDQ SRGEMVLVLY PAQDEKHEGL 201  SESAQNIMKI LTAELPTKQA AELAAKITGE GKKALYD..

Further work revealed the complete nucleotide sequence <SEQ ID 641>:

1 ATGTTTCAGA AACATTTGCA GAAAGCCTCC GACAGCGTCG TCGGAGGGAC 51 ATTATACGTGGTTGCCACGC CCATCGGCAA TTTGGCGGAC ATTACCCTGC 101 GCGCTTTGGC GGTATTGCAAAAGGCGGACA TCATCTGTGC CGAAGACACG 151 CGCGTTACCG CACAGCTTTT GAGCGCGTACGGCATTCAGG GCAAACTCGT 201 CAGTGTGCGC GAACACAACG AACGGCAGAT GGCGGACAAGATTGTCGGCT 251 ATCTTTCAGA CGGCATGGTT GTGGCACAGG TTTCCGATGC GGGTACGCCG301 GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC GTGAGGCCGG 351GTTTAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTGATG GCGGCTTTGA 401 GCGTGGCCGGTGTGGAAGGA TCCGATTTTT ATTTCAACGG TTTTGTACCG 451 CCGAAATCGG GAGAACGCAGGAAACTGTTT GCCAAATGGG TGCGGGCGGC 501 GTTTCCTATC GTCATGTTTG AAACGCCGCACCGCATCGGT GCGACGCTTG 551 CCGATATGGC GGAACTGTTC CCCGAACGCC GATTAATGCTGGCGCGCGAA 601 ATTACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG GGGAAATTCA651 GACGGCATTG TCTGCCGACG GCAACCAATC GCGCGGCGAG ATGGTGTTGG 701TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC CGAGTCCGCG 751 CAAAACATCATGAAAATCCT CACAGCCGAG CTGCCGACCA AACAGGCGGC 801 GGAGCTTGCT GCCAAAATCACGGGCGAGGG AAAGAAAGCT TTGTACGATC 851 TGGCTCTGTC TTGGAAAAAC AAATAG

This corresponds to the amino acid sequence <SEQ ID 642; ORF147-1>:

1 MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ KADIICAEDT 51 RVTAQLLSAYGIQGKLVSVR EHNERQMADK IVGYLSDGMV VAQVSDAGTP 101 AVCDPGAKLARRVREAGFKV VPVVGASAVM AALSVAGVEG SDFYFNGFVP 151 PKSGERRKLF AKWVRAAFPIVMFETPHRIG ATLADMAELF PERRLMLARE 201 ITKTFETFLS GTVGEIQTAL SADGNQSRGEMVLVLYPAQD EKHEGLSESA 251 QNIMKILTAE LPTKQAAELA AKITGEGKKA LYDLALSWKN K*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical Protein ORF286 of E. coli (Accession Number U18997)

ORF147 and E. coli ORF286 protein show 36% aa identity in 237aa overlap:

Orf147: 1 AEDTRVTAQLLSAYGIQGKLVSVREHNERQMADKIVGYLSDGMVVAQVSDAGTPAVCDPG60 AEDTR T  LL  +GI  +L ++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG Orf286:43 AEDTRHTGLLLQHFGINARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPG 102Orf147: 61 AKLARRVREXXXXXXXXXXXXXXXXXXXXXXXEGSDFYFNGFVPPKSGERRKLFAKWVRA120   L R  RE                           F + GF+P KS  RR Orf286: 103YHLVRTCREAGIRVVPLPGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAE 162 Orf147:121 AFPIVMFETPHRIGAALADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALSADGD 179   ++ +E+ HR+  +L D+  +  E R ++LARE+TKT+ET     VGE+   +  D + Orf286: 163PRTLIFYESTHRLLDSLEDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDEN 222 Orf147:180 QSRGEMVLVLYPAQDEKHEGLSESAQNIMKILTAELPTKQAAELAAKITGEGKKALY 236+ +GEMVL++      + E L   A   + +L AELP K+AA LAA+I G  K ALY Orf286: 223RRKGEMVLIV-EGHKAQEEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALY 278Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF147 shows 96.6% identity over a 237aa overlap with ORF75a from strainA of N. meningitidis:

ORF147a is identical to ORF75a, which includes aa 56-292 of ORF75.

Homology with a Predicted ORF from N. gonorrhoeae

ORF147 shows 94.1% identity over a 237aa overlap with a predicted ORF(ORF147ng) from N. gonorrhoeae:

An ORF147ng nucleotide sequence <SEQ ID 643> was predicted to encode aprotein having amino acid sequence <SEQ ID 644>:

1 MSVFQTAFFM FQKHLQKASD SVVGGTLYVV ATPIGNLADI TLRALAVLQK 51 ADIICAEDTRVTAQLLSAYG IQGRLVSVRE HNERQMADKV IGFLSDGLVV 101 AQVSDAGTPA VCDPGAKLARRVREAGFKVV PVVGASAVMA ALSVAGVAES 151 DFYFNGFVPP KSGERRKLFA KWVRAAFPVVMFETPHRIGA TLADMAELFP 201 ERRLMLAREI TKTFETFLSG TVGEIQTALA ADGNQSRGEMVLVLYPAQDE 251 KHEGLSESAQ NAMKILAAEL PTKQAAELAA KITGEGKKAL YDLALSWKNK301 *

Further work revealed the following gonococcal DNA sequence <SEQ ID645>:

1 ATGTTTCAGA AACACTTGCA GAAAGCCTCC GACAGCGTCG TCGGAGGGAC 51 ATTATACGTGGTTGCCACGC CCATCGGCAA TTTGGCAGAC ATTACCCTGC 101 GCGCTTTGGC GGTATTGCAAAAGGCGGACA TCATTTGTGC CGAAGACACG 151 CGCGTTACTG CGCAGCTTTT GAGCGCGTACGGCATTCAGG GCAGGTTGGT 201 CAGTGTGCGC GAACACAACG AGCGGCAGAT GGCGGACAAGGTAATCGGTT 251 TCCTTTCAGA CGGCCTGGTT GTGGCGCAGG TTTCCGATGC GGGTACGCCG301 GCCGTGTGCG ACCCGGGCGC GAAACTCGCC CGCCGCGTGC GCGAAGCAGG 351GTTCAAAGTC GTTCCCGTCG TGGGCGCAAG CGCGGTAATG GCGGCGTTGA 401 GTGTGGCCGGTGTGGCGGAA TCCGATTTTT ATTTCAACGG TTTTGTACCG 451 CCGAAATCGG GCGAACGTAGGAAATTGTTT GCCAAATGGG TGCGGGCGGC 501 ATTTCCTGTC GTCATGTTTG AAACGCCGCACCGAATCGGG GCAACGCTTG 551 CCGATATGGC GGAATTGTTC CCCGAACGCC GTCTGATGCTGGCGCGCGAA 601 ATCACGAAAA CGTTTGAAAC GTTCTTAAGC GGCACGGTTG GGGAAATTCA651 GACGGCATTG GCGGCGGACG GCAACCAATC GCGCGGCGAG ATGGTGTTGG 701TGCTTTATCC GGCGCAGGAT GAAAAACACG AAGGCTTGTC CGAGTCTGCG 751 CAAAATGCGATGAAAATCCT TGCGGCCGAG CTGCCGACCA AGCAGGCGGC 801 GGAGCTTGCC GCCAAGATTACAGGTGAGGG CAAAAAGGCT TTGTACGATT 851 TGGCACTGTC GTGGAAAAAC AAATGA

This corresponds to the amino acid sequence <SEQ ID 646; ORF147ng-1>:

1 MFQKHLQKAS DSVVGGTLYV VATPIGNLAD ITLRALAVLQ KADIICAEDT 51 RVTAQLLSAYGIQGRLVSVR EHNERQMADK VIGFLSDGLV VAQVSDAGTP 101 AVCDPGAKLARRVREAGFKV VPVVGASAVM AALSVAGVAE SDFYFNGFVP 151 PKSGERRKLF AKWVRAAFPVVMFETPHRIG ATLADMAELF PERRLMLARE 201 ITKTFETFLS GTVGEIQTAL AADGNQSRGEMVLVLYPAQD EKHEGLSESA 251 QNAMKILAAE LPTKQAAELA AKITGEGKKA LYDLALSWKN K*

ORF147ng shows homology to a hypothetical E. coli protein:

sp|P45528|YRAL_ECOLI HYPOTHETICAL 31.3 KD PROTEIN IN AGAI-MTR INTERGENICREGION (F286) >gi|606086 (U18997) ORF_f286 [Escherichiacoli] >gi|1789535 (AE000395) hypothetical 31.3 kD protein in agai-mtrintergenic region [Escherichia coli] Length = 286 Score = 218 bits(550), Expect = 3e−56 Identities = 128/284 (45%), Positives = 171/284(60%), Gaps = 4/284 (1%) Query: 4KHLQKASDSVVGGTLYVVATPIGNLADITLRALAVLQKADIICAEDTRVTAQLLSAYGIQ 63K  Q A +S   G LY+V TPIGNLADIT RAL VLQ  D+I AEDTR T  LL  +GI Sbjct: 2KQHQSADNSQ--GQLYIVPTPIGNLADITQRALEVLQAVDLIAAEDTRHTGLLLQHFGIN 59 Query:64 GRLVSVREHNERQMADKVIGFLSDGLVVAQVSDAGTPAVCDPGAKLARRVREAGFKVVPV 123 RL ++ +HNE+Q A+ ++  L +G  +A VSDAGTP + DPG  L R  REAG +VVP+ Sbjct: 60ARLFALHDHNEQQKAETLLAKLQEGQNIALVSDAGTPLINDPGYHLVRTCREAGIRVVPL 119 Query:124 VGASAVMAALSVAGVAESDFYFNGFVPPKSGERRKLFAKWVRAAFPVVMFETPHRIGATL 183 G  A + ALS AG+    F + GF+P KS  RR            ++ +E+ HR+  +L Sbjct: 120PGPCAAITALSAAGLPSDRFCYEGFLPAKSKGRRDALKAIEAEPRTLIFYESTHRLLDSL 179 Query:184 ADMAELFPERR-LMLAREITKTFETFLSGTVGEIQTALAADGNQSRGEMVLVLYPAQDEK 242 D+  +  E R ++LARE+TKT+ET     VGE+   +  D N+ +GEMVL++      + Sbjct: 180EDIVAVLGESRYVVLARELTKTWETIHGAPVGELLAWVKEDENRRKGEMVLIV-EGHKAQ 238 Query:243 HEGLSESAQNAMKILAAELPTKQAAELAAKITGEGKKALYDLAL 286 E L   A   + +L AELP K+AA LAA+I G  K ALY  AL Sbjct: 239EEDLPADALRTLALLQAELPLKKAAALAAEIHGVKKNALYKYAL 282

Based on the computer analysis and the presence of a putativetransmembrane domain in the gonococcal protein, it is predicted thatthese proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 77

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 647>

This corresponds to the amino acid sequence <SEQ ID 648; ORF1>:

Further sequencing analysis revealed the complete nucleotide sequence<SEQ ID 649>:

1 ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA AAGCCCCGAA 51 AACCGGCCGCATCCGCTTCT CGCCTGCTTA CTTAGCCATA TGCCTGTCGT 101 TCGGCATTCT TCCCCAAGCCTGGGCGGGAC ACACTTATTT CGGCATCAAC 151 TACCAATACT ATCGCGACTT TGCCGAAAATAAAGGCAAGT TTGCAGTCGG 201 GGCGAAAGAT ATTGAGGTTT ACAACAAAAA AGGGGAGTTGGTCGGCAAAT 251 CAATGACAAA AGCCCCGATG ATTGATTTTT CTGTGGTGTC GCGTAACGGC301 GTGGCGGCAT TGGTGGGCGA TCAATATATT GTGAGCGTGG CACATAACGG 351CGGCTATAAC AACGTTGATT TTGGTGCGGA AGGAAGAAAT CCCGATCAAC 401 ATCGTTTTACTTATAAAATT GTGAAACGGA ATAATTATAA AGCAGGGACT 451 AAAGGCCATC CTTATGGCGGCGATTATCAT ATGCCGCGTT TGCATAAATT 501 TGTCACAGAT GCAGAACCTG TTGAAATGACCAGTTATATG GATGGGCGGA 551 AATATATCGA TCAAAATAAT TACCCTGACC GTGTTCGTATTGGGGCAGGC 601 AGGCAATATT GGCGATCTGA TGAAGATGAG CCCAATAACC GCGAAAGTTC651 ATATCATATT GCAAGTGCGT ATTCTTGGCT CGTTGGTGGC AATACCTTTG 701CACAAAATGG ATCAGGTGGT GGCACAGTCA ACTTAGGTAG TGAAAAAATT 751 AAACATAGCCCATATGGTTT TTTACCAACA GGAGGCTCAT TTGGCGACAG 801 TGGCTCACCA ATGTTTATCTATGATGCCCA AAAGCAAAAG TGGTTAATTA 851 ATGGGGTATT GCAAACGGGC AACCCCTATATAGGAAAAAG CAATGGCTTC 901 CAGCTGGTTC GTAAAGATTG GTTCTATGAT GAAATCTTTGCTGGAGATAC 951 CCATTCAGTA TTCTACGAAC CACGTCAAAA TGGGAAATAC TCTTTTAACG1001 ACGATAATAA TGGCACAGGA AAAATCAATG CCAAACATGA ACACAATTCT 1051CTGCCTAATA GATTAAAAAC ACGAACCGTT CAATTGTTTA ATGTTTCTTT 1101 ATCCGAGACAGCAAGAGAAC CTGTTTATCA TGCTGCAGGT GGTGTCAACA 1151 GTTATCGACC CAGACTGAATAATGGAGAAA ATATTTCCTT TATTGACGAA 1201 GGAAAAGGCG AATTGATACT TACCAGCAACATCAATCAAG GTGCTGGAGG 1251 ATTATATTTC CAAGGAGATT TTACGGTCTC GCCTGAAAATAACGAAACTT 1301 GGCAAGGCGC GGGCGTTCAT ATCAGTGAAG ACAGTACCGT TACTTGGAAA1351 GTAAACGGCG TGGCAAACGA CCGCCTGTCC AAAATCGGCA AAGGCACGCT 1401GCACGTTCAA GCCAAAGGGG AAAACCAAGG CTCGATCAGC GTGGGCGACG 1451 GTACAGTCATTTTGGATCAG CAGGCAGACG ATAAAGGCAA AAAACAAGCC 1501 TTTAGTGAAA TCGGCTTGGTCAGCGGCAGG GGTACGGTGC AACTGAATGC 1551 CGATAATCAG TTCAACCCCG ACAAACTCTATTTCGGCTTT CGCGGCGGAC 1601 GTTTGGATTT AAACGGGCAT TCGCTTTCGT TCCACCGTATTCAAAATACC 1651 GATGAAGGGG CGATGATTGT CAACCACAAT CAAGACAAAG AATCCACCGT1701 TACCATTACA GGCAATAAAG ATATTGCTAC AACCGGCAAT AACAACAGCT 1751TGGATAGCAA AAAAGAAATT GCCTACAACG GTTGGTTTGG CGAGAAAGAT 1801 ACGACCAAAACGAACGGGCG GCTCAACCTT GTTTACCAGC CCGCCGCAGA 1851 AGACCGCACC CTGCTGCTTTCCGGCGGAAC AAATTTAAAC GGCAACATCA 1901 CGCAAACAAA CGGCAAACTG TTTTTCAGCGGCAGACCAAC ACCGCACGCC 1951 TACAATCATT TAAACGACCA TTGGTCGCAA AAAGAGGGCATTCCTCGCGG 2001 GGAAATCGTG TGGGACAACG ACTGGATCAA CCGCACATTT AAAGCGGAAA2051 ACTTCCAAAT TAAAGGCGGA CAGGCGGTGG TTTCCCGCAA TGTTGCCAAA 2101GTGAAAGGCG ATTGGCATTT GAGCAATCAC GCCCAAGCAG TTTTTGGTGT 2151 CGCACCGCATCAAAGCCACA CAATCTGTAC ACGTTCGGAC TGGACGGGTC 2201 TGACAAATTG TGTCGAAAAAACCATTACCG ACGATAAAGT GATTGCTTCA 2251 TTGACTAAGA CCGACATCAG CGGCAATGTCGATCTTGCCG ATCACGCTCA 2301 TTTAAATCTC ACAGGGCTTG CCACACTCAA CGGCAATCTTAGTGCAAATG 2351 GCGATACACG TTATACAGTC AGCCACAACG CCACCCAAAA CGGCAACCTT2401 AGCCTCGTGG GCAATGCCCA AGCAACATTT AATCAAGCCA CATTAAACGG 2451CAACACATCG GCTTCGGGCA ATGCTTCATT TAATCTAAGC GACCACGCCG 2501 TACAAAACGGCAGTCTGACG CTTTCCGGCA ACGCTAAGGC AAACGTAAGC 2551 CATTCCGCAC TCAACGGTAATGTCTCCCTA GCCGATAAGG CAGTATTCCA 2601 TTTTGAAAGC AGCCGCTTTA CCGGACAAATCAGCGGCGGC AAGGATACGG 2651 CATTACACTT AAAAGACAGC GAATGGACGC TGCCGTCAGGCACGGAATTA 2701 GGCAATTTAA ACCTTGACAA CGCCACCATT ACACTCAATT CCGCCTATCG2751 CCACGATGCG GCAGGGGCGC AAACCGGCAG TGCGACAGAT GCGCCGCGCC 2801GCCGTTCGCG CCGTTCGCGC CGTTCCCTAT TATCCGTTAC ACCGCCAACT 2851 TCGGTAGAATCCCGTTTCAA CACGCTGACG GTAAACGGCA AATTGAACGG 2901 TCAGGGAACA TTCCGCTTTATGTCGGAACT CTTCGGCTAC CGCAGCGACA 2951 AATTGAAGCT GGCGGAAAGT TCCGAAGGCACTTACACCTT GGCGGTCAAC 3001 AATACCGGCA ACGAACCTGC AAGCCTCGAA CAATTGACGGTAGTGGAAGG 3051 AAAAGACAAC AAACCGCTGT CCGAAAACCT TAATTTCACC CTGCAAAACG3101 AACACGTCGA TGCCGGCGCG TGGCGTTACC AACTCATCCG CAAAGACGGC 3151GAGTTCCGCC TGCATAATCC GGTCAAAGAA CAAGAGCTTT CCGACAAACT 3201 CGGCAAGGCAGAAGCCAAAA AACAGGCGGA AAAAGACAAC GCGCAAAGCC 3251 TTGACGCGCT GATTGCGGCCGGGCGCGATG CCGTCGAAAA GACAGAAAGC 3301 GTTGCCGAAC CGGCCCGGCA GGCAGGCGGGGAAAATGTCG GCATTATGCA 3351 GGCGGAGGAA GAGAAAAAAC GGGTGCAGGC GGATAAAGACACCGCCTTGG 3401 CGAAACAGCG CGAAGCGGAA ACCCGGCCGG CTACCACCGC CTTCCCCCGC3451 GCCCGCCGCG CCCGCCGGGA TTTGCCGCAA CTGCAACCCC AACCGCAGCC 3501CCAACCGCAG CGCGACCTGA TCAGCCGTTA TGCCAATAGC GGTTTGAGTG 3551 AATTTTCCGCCACGCTCAAC AGCGTTTTCG CCGTACAGGA CGAATTAGAC 3601 CGCGTATTTG CCGAAGACCGCCGCAACGCC GTTTGGACAA GCGGCATCCG 3651 GGACACCAAA CACTACCGTT CGCAAGATTTCCGCGCCTAC CGCCAACAAA 3701 CCGACCTGCG CCAAATCGGT ATGCAGAAAA ACCTCGGCAGCGGGCGCGTC 3751 GGCATCCTGT TTTCGCACAA CCGGACCGAA AACACCTTCG ACGACGGCAT3801 CGGCAACTCG GCACGGCTTG CCCACGGCGC CGTTTTCGGG CAATACGGCA 3851TCGACAGGTT CTACATCGGC ATCAGCGCGG GCGCGGGTTT TAGCAGCGGC 3901 AGCCTTTCAGACGGCATCGG AGGCAAAATC CGCCGCCGCG TGCTGCATTA 3951 CGGCATTCAG GCACGATACCGCGCCGGTTT CGGCGGATTC GGCATCGAAC 4001 CGCACATCGG CGCAACGCGC TATTTCGTCCAAAAAGCGGA TTACCGCTAC 4051 GAAAACGTCA ATATCGCCAC CCCCGGCCTT GCATTCAACCGCTACCGCGC 4101 GGGCATTAAG GCAGATTATT CATTCAAACC GGCGCAACAC ATTTCCATCA4151 CGCCTTATTT GAGCCTGTCC TATACCGATG CCGCTTCGGG CAAAGTCCGA 4201ACACGCGTCA ATACCGCCGT ATTGGCTCAG GATTTCGGCA AAACCCGCAG 4251 TGCGGAATGGGGCGTAAACG CCGAAATCAA AGGTTTCACG CTGTCCCTCC 4301 ACGCTGCCGC CGCCAAAGGCCCGCAACTGG AAGCGCAACA CAGCGCGGGC 4351 ATCAAATTAG GCTACCGCTG GTAA

This corresponds to the amino acid sequence <SEQ ID 650; ORF1-1>:

1 MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA WAGHTYFGIN 51 YQYYRDFAENKGKFAVGAKD IEVYNKKGEL VGKSMTKAPM IDFSVVSRNG 101 VAALVGDQYI VSVAHNGGYNNVDFGAEGRN PDQHRFTYKI VKRNNYKAGT 151 KGHPYGGDYH MPRLHKFVTD AEPVEMTSYMDGRKYIDQNN YPDRVRIGAG 201 RQYWRSDEDE PNNRESSYHI ASAYSWLVGG NTFAQNGSGGGTVNLGSEKI 251 KHSPYGFLPT GGSFGDSGSP MFIYDAQKQK WLINGVLQTG NPYIGKSNGF301 QLVRKDWFYD EIFAGDTHSV FYEPRQNGKY SFNDDNNGTG KINAKHEHNS 351LPNRLKTRTV QLFNVSLSET AREPVYHAAG GVNSYRPRLN NGENISFIDE 401 GKGELILTSNINQGAGGLYF QGDFTVSPEN NETWQGAGVH ISEDSTVTWK 451 VNGVANDRLS KIGKGTLHVQAKGENQGSIS VGDGTVILDQ QADDKGKKQA 501 FSEIGLVSGR GTVQLNADNQ FNPDKLYFGFRGGRLDLNGH SLSFHRIQNT 551 DEGAMIVNHN QDKESTVTIT GNKDIATTGN NNSLDSKKEIAYNGWFGEKD 601 TTKTNGRLNL VYQPAAEDRT LLLSGGTNLN GNITQTNGKL FFSGRPTPHA651 YNHLNDHWSQ KEGIPRGEIV WDNDWINRTF KAENFQIKGG QAVVSRNVAK 701VKGDWHLSNH AQAVFGVAPH QSHTICTRSD WTGLTNCVEK TITDDKVIAS 751 LTKTDISGNVDLADHAHLNL TGLATLNGNL SANGDTRYTV SHNATQNGNL 801 SLVGNAQATF NQATLNGNTSASGNASFNLS DHAVQNGSLT LSGNAKANVS 851 HSALNGNVSL ADKAVFHFES SRFTGQISGGKDTALHLKDS EWTLPSGTEL 901 GNLNLDNATI TLNSAYRHDA AGAQTGSATD APRRRSRRSRRSLLSVTPPT 951 SVESRFNTLT VNGKLNGQGT FRFMSELFGY RSDKLKLAES SEGTYTLAVN1001 NTGNEPASLE QLTVVEGKDN KPLSENLNFT LQNEHVDAGA WRYQLIRKDG 1051EFRLHNPVKE QELSDKLGKA EAKKQAEKDN AQSLDALIAA GRDAVEKTES 1101 VAEPARQAGGENVGIMQAEE EKKRVQADKD TALAKQREAE TRPATTAFPR 1151 ARRARRDLPQ LQPQPQPQPQRDLISRYANS GLSEFSATLN SVFAVQDELD 1201 RVFAEDRRNA VWTSGIRDTK HYRSQDFRAYRQQTDLRQIG MQKNLGSGRV 1251 GILFSHNRTE NTFDDGIGNS ARLAHGAVFG QYGIDRFYIGISAGAGFSSG 1301 SLSDGIGGKI RRRVLHYGIQ ARYRAGFGGF GIEPHIGATR YFVQKADYRY1351 ENVNIATPGL AFNRYRAGIK ADYSFKPAQH ISITPYLSLS YTDAASGKVR 1401TRVNTAVLAQ DFGKTRSAEW GVNAEIKGFT LSLHAAAAKG PQLEAQHSAG 1451 IKLGYRW*

Computer analysis of these sequences gave the following results:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF1 shows 57.8% identity over a 1456aa overlap with an ORF (ORF1a) fromstrain A of N. meningitidis:

The complete length ORF1a nucleotide sequence <SEQ ID 651> is:

1 ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA AAGCCCCGAA 51 AACCGGCCGCATCCGCTTCT CGCCTGCTTA CTTAGCCATA TGCCTGTCGT 101 TCGGCATTCT TCCCCAAGCTTGGGCGGGAC ACACTTATTT CGGCATCAAC 151 TACCAATACT ATCGCGACTT TGCCGAAAATAAAGGCAAGT TTGCAGTCGG 201 GGCGAAAGAT ATTGAGGTNT ACAACAAAAA AGGGGAGTTGGTCGGCAAAT 251 CAATGACAAA AGCCCCGATG ATTGATTTTT CTGTGGTGTC GCGTAACGGC301 GTGGCGGCAT TGGTGGGCGA TCAATATATT GTGAGCGTGG CACATAACGG 351CGGCTATAAC AACGTTGATT TTGGTGCGGA AGGAAGNAAT CCCGATCAGC 401 ACCGTTTTTCTTACCAAATT GTGAAAAGAA ATAATTATAA GCCTGACAAT 451 TCACACCCTT ACAACGGCGATTANCATATG CCGCGTTTGC ATAAATTTGT 501 CACAGATGCA GAACCTGTCG AAATGACGAGTGACATGAGG GGGAATACCT 551 ATTCCGATAA AGAAAAATAT CCCGAGCGTG TCCGCATCGGCTCAGGACAC 601 CACTATTGGC GTTATGATGA TGACAAACAC GGCGATTTAT CCTACTCCGG651 CGCATGGTTA ATTGGCGGCA ATACACATAT GCAGGGTTGG GGAAATAATG 701GCGTANTTAG TTTGAGCGGC GATGTGCGCC ATGCCAACGA CTATGGCCCT 751 ATGCCGATTGCAGGTGCGGC AGGCGACAGC GGTTCGCCAA TGTTTATTTA 801 TGACAAAACA AACAATAAATGGCTGCTCAA CGGAGTTTTA CAAACCGGCT 851 ACCCTTATTC CGGCAGGGAA AACGGTTTCCAGCTGATACG CAAAGATTGG 901 TTCTACGATG ACATTTACAG AGGCGATACA CATACCGTCTNTTTTGAACC 951 GCGCAGTAAC GGACATTTTT CCTTTACATC CAACAACAAC GGTACGGGTA1001 CGGTAACAGA AACCAACGAA AAGGTNTCCA ATCCAAAGCT TAAAGTACAG 1051ACAGTCCGAC TGTTTGACGA ATCTTTGAAT GAAACTGATA AAGAACCAGT 1101 TTACGCGGCAGGGGGTGTTA ATCAGTACCG TCCAAGGTTA AACAACGGTG 1151 AAAACCTTTC TTTTATCGATTACGGCAACG GCAAACTCAT CTTATCAAAC 1201 AACATCAACC AAGGCGCGGG CGGTTTGTATTTTGAAGGTG ATTTTACGGT 1251 CTCGCCTGAA AACAACGAAA CGTGGCAAGG CGCGGGCGTTCATATCAGTG 1301 AAGACAGTAC CGTTACTTGG AAAGTAAACG GCGTGGCAAA CGACCGCCTG1351 TCCAAAATCG GCAAAGGCAC GCTGCACGTT CAAGCCAAAG GGGAAAACCA 1401AGGCTCGATC AGCGTGGGCG ACGGTACAGT CATTTTGGAT CAGCAGGCAG 1451 ACGATAAAGGCAAAAAACAA GCCTTTAGTG AAATCGGCTT GNTCAGCGGC 1501 AGGGGTACGG TGCAACTGAATGCCGATAAT CAGTTCAACC CCGACAAACT 1551 CTATTTCGGC TTTCGCGGCG GACGTTTGGATTTAAACGGG CATTCGCTTT 1601 CGTTCCACCG TATTCAAAAT ACCGATGAAG GGGCGATGATTGNCNATCAT 1651 AATGCCACAA CAACATCCAC CGTTACCATT ACAGGGAATG AAAGTATTAC1701 ACAACCGAGT GGTAAGAATA TCAATAGACT TAATTACAGC AAAGAAATTG 1751CCTACAACGG TTGGTTTGGC GAGAAAGATA CGACCAAAAC GAACGGGCGG 1801 CTCAACCTTGTTTACCAGCC CGCCGCAGAA GACCGCACCC NGCTGCTTTC 1851 CGGCGGAACA AATTTAAACGGCAACATCAC GCAAACAAAC GGCAAACTGT 1901 TTTTCAGCGG CAGACCGACA CCGCACGCCTACAATCATTT AGGAAGCGGG 1951 TGGTCAAAAA TGGAAGGTAT CCCACAAGGA GAAATCGTGTGGGACAACGA 2001 CTGGATCNAC CGCACGTTTA AAGCGGAAAA TTTCCATATT CAGGGCGGGC2051 AGGCGGTGAT TTCCCGCAAT GTTGCCAAAG TGGAAGGCGA TTGNCATTTG 2101AGCAATCACG CCCAAGCAGT TTTTGGTGTC GCACCGCATC AAAGCCATAC 2151 AATCTGTACACGTTCGGACT GGACNGGTCT GACAAATTGT GTCGAANAAA 2201 NCATTACCGA CGATAAAGTGATTGCTTCAT TGACTAAGAC NGACNTNAGC 2251 GGCANTGTNA GNCTNNCCNA TNACGNTNNTTNAAANCTCN CNGGGCNTGC 2301 NNCACTNAAN GGCAATCTTA GTGCAAATGG CGATACACGTTATACAGTCA 2351 GCCACAACGC CACCCAAAAC GGCAACCTTA GCCTCGTGGG CAATGCCCAA2401 GCAACATTTA ATCAAGCCAC ATTAAACGGC AACNCATCGG NTTCGGGCAA 2451TGCTTCATTT AATCTAAGCA ACAACGCCGC ACAAAACGGC AGTCTGACGC 2501 TTTCCGACAACGCTAAGGCA AACGTAAGCC ATTCCGCACT CAACGGCAAT 2551 GTCTCCCTAG CCGATAAGGCAGTATTCCAT TTTGAAAACA GCCGCTTTAC 2601 CGGACAACTC AGCGGCAGCA AGGANACAGCATTACACTTA AAAGACAGCG 2651 AATGGACGCT GCCGTCAGGC ACGGAATTAG GCAATTTAAACCTTGACAAC 2701 GCCACCATTA CACTCAATTC CGCCTATCGC CACGATGCTG CAGGCGCGCA2751 AACCGGCAGN GTGTCAGACA CGCCGCGCCG CCGTTCGCGC CGTTCCCTAT 2801TATCCGTTAC ACCGCCAACT TCGGTAGAAT CCCGTTTCAA CACGCTGACG 2851 GTAAACGGCAAATTGAACNG TCAAGGAACA TTCCGCTTTA TGTCGGAACT 2901 CTTCGGCTAC CGAAGCGACAAATTGAAGCT GGCGGAAAGT TCCGAAGGNA 2951 CTTACACCTT GGCGGTCAAC AATACCGGCAACGAACCCGT AAGCCTCGAT 3001 CAATTGACGG TAGTGGAAGG GAAAGACAAC AAACCGCTGTCCGAAAACCT 3051 TAATTTCACC CTGCAAAACG AACACGTCGA TGCCGGCGCG TGGCGTTACC3101 AACTCATCCG CAAAGACGGC GAGTTCCGCC TGCATAATCC GGTCAAAGAA 3151CAAGAGCTTT CCGACAAACT CGGCAAGGCA GAAGCCAAAA AACAGGCGGA 3201 AAAAGACAACGCGCAAAGCC TTGACGCGCT GATTGCGGCC GGGCGCGATG 3251 CCGCCGAAAA GACAGAAAGCGTTGCCGAAC CGGCCCGGCN GGCAGGCGGG 3301 GAAAATGTCG GCATTATGCA GGCGGAGGAAGAGAAAAAAC GGGTGCAGGC 3351 GGATAAAGAC AGCGCNTTGG CGAAACAGCG CGAAGCGGAAACCCGGCCGG 3401 NTACCACCGC CTTCCCCCGC GCCCGCNGCG CCCGCCGGGA TTTGCCGCAA3451 CCGCAGCCCC AACCGCAACC TCAACCCCAA CCGCAGCGCG ACCTGATNAG 3501CCGTTATGCC AATAGCGGTT TGAGTGAATT TTCCGCCACG CTCAACAGCG 3551 TTTTCGCCGTACAGGACGAA TTGGACCGCG TGTTTGCCGA AGACCGCCGC 3601 AACGCNGTTT GGACAAGCNGCATCCGGNAC ACCAAACACT ACCGTTCGCA 3651 AGATTTCCGC GCCTACCGCC AACAAACCGACCTGCGCCAA ATCGGTATGC 3701 AGAAAAACCT CGGCAGCGGG CGCGTCGGCA TCCTGTTTTCGCACAACCGG 3751 ACCGAAAACA NCTTCGACGA CGGCATCGGC AACTCGGCAC GGCTTGCCCA3801 CGGCGCCGTT TTCGGGCAAT ACGGCATCGG CAGGTTCGAC ATCGGCATCA 3851GCACGGGCGC GGGTTTTAGC AGCGGCANTC TNTCAGACGG CATCGGAGGC 3901 AAAATCCGCCGCCGCGTGCT GCATTACGGC ATTCAGGCAC GATACCGCGC 3951 CGGTTTCGGC GGATTCGGCATCGAACCGTA CATCGGCGCA ACGCGCTATT 4001 TCGTCCAAAA AGCGGATTAC CGCTACGAAAACGTCAATAT CGCCACCCCC 4051 GGTCTTGCGT TCAACCGNTA CCGNGCGGGC ATTAAGGCAGATTATTCATT 4101 CAAACCGGCG CAACACATNT CCATCACNCC TTATTTNAGC CTGTCCTATA4151 CCGATGCCGC TTCGGGCAAA GTCCGAACAC GCGTCAATAC CGCNGTATTG 4201GCTCAGGATT TCGGCAAAAC CCGCAGTGCG GAATGGGGCG TAAACGCCGA 4251 AATCAAAGGTTTCACGCTGT CCNTCCACGC TGCCGCCGCC AAAGGNCCGC 4301 AACTGGAAGC GCAACACAGCGCGGGCATCA AATTAGGCTA CCGCTGGTAA

This encodes a protein having amino acid sequence <SEQ ID 652>:

1 MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA WAGHTYFGIN 51 YQYYRDFAENKGKFAVGAKD IEVYNKKGEL VGKSMTKAPM IDFSVVSRNG 101 VAALVGDQYI VSVAHNGGYNNVDFGAEGXN PDQHRFSYQI VKRNNYKPDN 151 SHPYNGDXHM PRLHKFVTDA EPVEMTSDMRGNTYSDKEKY PERVRIGSGH 201 HYWRYDDDKH GDLSYSGAWL IGGNTHMQGW GNNGVXSLSGDVRHANDYGP 251 MPIAGAAGDS GSPMFIYDKT NNKWLLNGVL QTGYPYSGRE NGFQLIRKDW301 FYDDIYRGDT HTVXFEPRSN GHFSFTSNNN GTGTVTETNE KVSNPKLKVQ 351TVRLFDESLN ETDKEPVYAA GGVNQYRPRL NNGENLSFID YGNGKLILSN 401 NINQGAGGLYFEGDFTVSPE NNETWQGAGV HISEDSTVTW KVNGVANDRL 451 SKIGKGTLHV QAKGENQGSISVGDGTVILD QQADDKGKKQ AFSEIGLXSG 501 RGTVQLNADN QFNPDKLYFG FRGGRLDLNGHSLSFHRIQN TDEGAMIXXH 551 NATTTSTVTI TGNESITQPS GKNINRLNYS KEIAYNGWFGEKDTTKTNGR 601 LNLVYQPAAE DRTXLLSGGT NLNGNITQTN GKLFFSGRPT PHAYNHLGSG651 WSKMEGIPQG EIVWDNDWIX RTFKAENFHI QGGQAVISRN VAKVEGDXHL 701SNHAQAVFGV APHQSHTICT RSDWTGLTNC VEXXITDDKV IASLTKTDXS 751 GXVXLXXXXXXXLXGXAXLX GNLSANGDTR YTVSHNATQN GNLSLVGNAQ 801 ATFNQATLNG NXSXSGNASFNLSNNAAQNG SLTLSDNAKA NVSHSALNGN 851 VSLADKAVFH FENSRFTGQL SGSKXTALHLKDSEWTLPSG TELGNLNLDN 901 ATITLNSAYR HDAAGAQTGX VSDTPRRRSR RSLLSVTPPTSVESRFNTLT 951 VNGKLNXQGT FRFMSELFGY RSDKLKLAES SEGTYTLAVN NTGNEPVSLD1001 QLTVVEGKDN KPLSENLNFT LQNEHVDAGA WRYQLIRKDG EFRLHNPVKE 1051QELSDKLGKA EAKKQAEKDN AQSLDALIAA GRDAAEKTES VAEPARXAGG 1101 ENVGIMQAEEEKKRVQADKD SALAKQREAE TRPXTTAFPR ARXARRDLPQ 1151 PQPQPQPQPQ PQRDLXSRYANSGLSEFSAT LNSVFAVQDE LDRVFAEDRR 1201 NAVWTSXIRX TKHYRSQDFR AYRQQTDLRQIGMQKNLGSG RVGILFSHNR 1251 TENXFDDGIG NSARLAHGAV FGQYGIGRFD IGISTGAGFSSGXLSDGIGG 1301 KIRRRVLHYG IQARYRAGFG GFGIEPYIGA TRYFVQKADY RYENVNIATP1351 GLAFNRYRAG IKADYSFKPA QHXSITPYXS LSYTDAASGK VRTRVNTAVL 1401AQDFGKTRSA EWGVNAEIKG FTLSXHAAAA KGPQLEAQHS AGIKLGYRW*

A transmembrane region is underlined.

ORF1-1 shows 86.3% identity over a 1462aa overlap with ORF1a:

Homology with Adhesion and Penetration Protein Hap Precursor of H.influenzae (Accession Number P45387)

Amino acids 23-423 of ORF1 show 59% aa identity with hap protein in450aa overlap:

orf1 23 FXAAYLAICLSFGILPQAWAGHTYFGINYQYYRDFAENKGKFAVGAKDIEVYNKKGELVG 82F   +L  C+S GI  QAWAGHTYFGI+YQYYRDFAENKGKF VGAK+IEVYNK+G+LVG hap 6FRLNFLTACVSLGIASQAWAGHTYFGIDYQYYRDFAENKGKFTVGAKNIEVYNKEGQLVG 65 orf1 83KSMTKAPMIDFSVVSRNGVAALVGVQYIVSVAHNGGYNNVDFGAEGXNIXDQXRXTYKIV 142 SMTKAPMIDFSVVSRNGVAALVG QYIVSVAHNGGYN+VDFGAEG N  DQ R TY+IV hap 66TSMTKAPMIDFSVVSRNGVAALVGDQYIVSVAHNGGYNDVDFGAEGRN-PDQHRFTYQIV 124 orf1143 KRNNYKAGTKGHPYGGDYHMPRLHKXVTDAEPVEMTSYMDGRKYIDQNNYPDRVRIGAGR 202KRNNY+A  + HPY GDYHMPRLHK VT+AEPV MT+ MDG+ Y D+ NYP+RVRIG+GR hap 125KRNNYQAWERKHPYDGDYHMPRLHKFVTEAEPVGMTTNMDGKVYADRENYPERVRIGSGR 184 orf1203 QYWRSDEDEPNNRESSYHIA---------------------------------------- 222QYWR+D+DE  N  SSY+++ hap 185QYWRTDKDEETNVHSSYYVSGAYRYLTAGNTHTQSGNGNGTVNLSGNVVSPNHYGPLPTG 244 orf1223 -----SGSPMFIYDAQKQKWLINGVLQTGNPYIGKSNGFQLVRKDWFYDEIFAGDTHSVF 277     SGSPMFIYDA+K++WLIN VLQTG+P+ G+ NGFQL+R++WFY+E+ A DT SVF hap 245GSKGDSGSPMFIYDAKKKQWLINAVLQTGHPFFGRGNGFQLIREEWFYNEVLAVDTPSVF 304 orf1278 --YEPRQNGKYSFNDDNNGTGKIN-AKHEHNSLPNRLKTRTVQLFNVSLSETAREPVYHA 334   YP  NG YSF  +N+GTGK+   +   +    + +  TV+LFN SL++TA+E V  A hap 305QRYIPPINGHYSFVSNNDGTGKLTLTRPSKDGSKAKSEVGTVKLFNPSLNQTAKEHV-KA 363 orf1335 AGGVNSYRPRLNNGENISFIDEGKGELILTSNINQGAGGLYFQGDFTV-SPENNETWQGA 393 A GN Y+PR+  G+NI   D+GKG L + +NINQGAGGLYF+G+F V   +NN TWQGA hap 364AAGYNIYQPRMEYGKNIYLGDQGKGTLTIENNINQGAGGLYFEGNFVVKGKQNNITWQGA 423 orf1394 GVHISEDSTVTWKVNGVANDRLSKIGKGTL 423 GV I +D+TV WKV+   NDRLSKIG GTLhap 424 GVSIGQDATVEWKVHNPENDRLSKIGIGTL 453

Amino acids 715-1011 of ORF1 show 50% aa identity with hap protein in258aa overlap:

Orf1 41 DTRYTVSHNATQ-NGNXSLVXNAQATFNQ-ATLNGNTSASGNASFNLSDHAVQNGSLTLS 98DT+   S   TQ NG+ +L  NA    +  A LNGN +   ++ F LS++A Q G++ LS hap 733DTKVINSIPITQINGSINLTNNATVNIHGLAKLNGNVTLIDHSQFTLSNNATQTGNIKLS 792 orf1 99GNAKANVSHSALNGNVSLADKAVFHFESSRFTGQISGGKDTALHLKDSEWTLPSGXELGN 158  +A AV+++ LNGNV L D A F  ++S F  QI G KDT + L+++ WT+PS   L N hap 793NHANATVNNATLNGNVHLTDSAQFSLKNSHFWHQIQGDKDTTVTLENATWTMPSDTTLQN 852 orf1159 LNLDNATITLNSAYRHDAAGAQTGSATDAPXXXXXXXXXXLLXVTPPTSVESRFNTLTVN 218 LL+N+T+TLNSAY        + S+ +AP          L   T PTS E RFNTLTVN hap 853LTLNNSTVTLNSAY--------SASSNNAPRHRRS-----LETETTPTSAEHRFNTLTVN 899 orf1219 GKLNGQGTFRFMSELFGYRSDKLKLAESSEGTYTLAVNNTGNEPASLEQLTVVEGKDNKP 278GKL+GQGTF+F S LFGY+SDKLKL+  +EG YTL+V NTG EP +LEQLT++E  DNKP hap 900GKLSGQGTFQFTSSLFGYKSDKLKLSNDAEGDYTLSVRNTGKEPVTLEQLTLIESLDNKP 959 orf1279 LSENLNFTLQNEHVDAGA 296 LS+ L FTL+N+HVDAGA hap 960 LSDKLKFTLENDHVDAGA977

Amino acids 1192-1450 of ORF1 show 41% aa identity with hap protein in259aa overlap:

Orf1 1 LDRVFAEDRRNAVWTSGIRDTKHYRSQDFRAYRQQTDLRQIGMQKNLGSGRVGILFSHNR 60LDR+F +  ++AVWT+  +D + Y S  FRAY+Q+T+LRQIG+QK L +GR+G +FSH+R hap 1135LDRLFVDQAQSAVWTNIAQDKRRYDSDAFRAYQQKTNLRQIGVQKALANGRIGAVFSHSR 1194 orf161 TENTFDDGIGNSARLAHGAVFGQYGIDRFYXXXXXXXXXXXXXXXXXIGXKXRRRVLHYG 120++NTFD+ + N A L   + F QY                          K  R+ ++YG hap 1195SDNTFDEQVKNHATLTMMSGFAQYQWGDLQFGVNVGTGISASKMAEEQSRKIHRKAINYG 1254 orf1121 IQARYRAGFGGFGIEPHIGATRYFVQKADYRYENVNIATPGLAFNRYRAGIKADYSFKPA 180 + AY+   G  GI+P+ G  RYF+++ +Y+ E V + TP LAFNRY AGI+ DY+F P hap 1255VNASYQFRLGQLGIQPYFGVNRYFIERENYQSEEVRVKTPSLAFNRYNAGIRVDYTFTPT 1314 orf1181 QHISITPYLSLSYTDAASGKVRTRVNTAVLAQDFGKTRSAEWGVNAEIKGFTLSLHAAAA 240 +IS+ PY  ++Y D ++  V+T VN  VL Q FG+    E G+ AEI  F +S   + + hap 1315DNISVKPYFFVNYVDVSNANVQTTVNLTVLQQPFGRYWQKEVGLKAEILHFQISAFISKS 1374 orf1241 KGPQLEAQHSAGIKLGYRW 259 +G QL  Q + G+KLGYRW hap 1375QGSQLGKQQNVGVKLGYRW 1393Homology with a Predicted ORF from N. gonorrhoeae

The blocks of ORF1 show 83.5%, 88.3%, and 97.7% identities in 467, 298,and 259 aa overlap, respectively with a predicted ORF (ORF1ng) from N.gonorrhoeae:

The complete length ORF1ng nucleotide sequence was identified <SEQ ID653>:

1 ATGAAAACAA CCGACAAACG GACAACCGAA ACACACCGCA AAGCCCCTAA 51 AACCGGCCGCATCCGCTTCT CGCCCGCTTA CTTAGCCATA TGCCTGTCGT 101 TCGGCATTCT GCCCCAAGCCCGGGCGGGAC ACACTTATTT CGGCATCAAC 151 TACCAATACT ATCGCGACTT TGCCGAAAATAAAGGCAAGT TTGCAGTCGG 201 GGCGAAAGAT ATTGAGGTTT ACAACAAAAA AGGGGAGTTGGTCGGCAAAT 251 CGATGACGAA AGCCCCGATG ATTGATTTTT CTGTGGTATC GCGTAACGGC301 GTGGCGGCAT TGGCGGGCGA TCAATATATT GTGAGCGTGG CACATAACGG 351CGGCTATAAC AATGTTGATT TTGGTGCGGA GGGAAGCAAT CCCGATCAGC 401 ACCGCTTTTCTTACCAAATT GTGAAAAGAA ATAATTATAA AGCAGGGACT 451 AACGGCCATC CTTATGGCGGCGATTATCAT ATGCCGCGTT TGCACAAATT 501 TGTCACAGAT GCAGAACCTG TTGAGATGACCAGTTATATG GATGGGTGGA 551 AATACGCTGA TTTAAATAAA TACCCTGATC GTGTTCGAATCGGAGCAGGC 601 AGACAATATT GGCGGTCTGA TGAAGACGAA CCCAATAACC GCGAAAGTTC651 ATATCATATT GCAAGCGCAT ATTCTTGGCT CGTCGGTGGC AATACCTTTG 701CACAAAATGG ATCAGGTGGT GGCACAGTCA ACTTAGGTAG CGAAAAAATT 751 AAACATAGCCCATATGGTTT TTTACCAACA GGAGGCTCAT TTGGCGACAG 801 TGGCTCACCA ATGTTTATCTATGATGCCCA AAAGCAAAAG TGGTTAATTA 851 ATGGGGTATT GCAAACAGGC AACCCCTATATAGGAAAAAG CAATGGCTTC 901 CAGCTAGTTC GTAAAGATTG GTTCTATGAT GAAATCTTTGCTGGAGATAC 951 CCATTCAGTA TTCTACGAAC CACATCAAAA TGGGAAATAC TTTTTTAACG1001 ACAATAATAA TGGCGCAGGA AAAATCGATG CCAAACATAA ACACTATTCT 1051CTACCTTATA GATTAAAAAC ACGAACCGTT CAATTGTTTA ATGTTTCTTT 1101 ATCCGAGACAGCAAGAGAAC CTGTTTATCA TGCTGCAGGT GGGGTCAACA 1151 GTTATCGACC CAGACTGAATAATGGAGAAA ATATTTCCTT TATTGACAAA 1201 GGAAAAGGTG AATTGATACT TACCAGCAACATCAACCAAG GCGCGGGCGG 1251 TTTGTATTTT GAGGGTAATT TTACGGTCTC GCCTAAAAACAACGAAACGT 1301 GGCAAGGCGC GGGCGTTCAT ATCAGTGATG GCAGTACCGT TACTTGGAAA1351 GTAAACGGCG TGGCAAACGA CCGCCTGTCC AAAATCGGCA AAGGCACGCT 1401GCTGGTTCAA GCCAAAGGGG AAAACCAAGG CTCGGTCAGC GTGGGCGACG 1451 GTAAAGTCATCTTAGATCAG CAGGCGGACG ATCAAGGCAA AAAACAAGCC 1501 TTTAGTGAAA TCGGCTTGGTCAGCGGCAGG GGGACGGTGC AACTGAATGC 1551 CGATAATCAG TTCAACCCCG ACAAACTCTATTTCGGCTTT CGCGGCGGAC 1601 GTTTGGATTT GAACGGGCAT TCGCTTTCGT TCCACCGCATTCAAAATACC 1651 GATGAAGGGG CGATGATTGT CAACCACAAT CAAGACAAAG AATCCACCGT1701 TACCATTACA GGCAATAAAG ATATTACTAC AACCGGCAAT AACAACAACT 1751TGGATAGCAA AAAAGAAATT GCCTACAACG GTTGGTTTGG CGAGAAAGAT 1801 GCAACCAAAACGAACGGGCG GCTCAATCTG AATTACCAAC CGGAAGAAGC 1851 GGATCGCACT TTACTGCTTTCCGGCGGAAC AAATTTAAAC GGCAATATCA 1901 CGCAAACAAA CGGCAAACTG TTTTTCAGCGGCAGACCGAC ACCGCACGCC 1951 TACAATCATT TAGGAAGCGG GTGGTCAAAA ATGGAAGGTATCCCACAAGG 2001 AGAAATCGTG TGGGACAACG ATTGGATCGA CCGCACATTT AAAGCGGAAA2051 ACTTCCATAT TCAGGGCGGA CAAGCGGTGG TTTCCCGCAA TGTTGCCAAA 2101GTGGAAGGCG ATTGGCATTT AAGCAATCAC GCCCAAGCAG TTTTCGGTGT 2151 CGCACCGCATCAAAGCCACA CAATCTGTAC ACGTTCGGAC TGGACGGGTC 2201 TGACAAGTTG TACCGAAAAAACCATTACCG ACGATAAAGT GATTGCTTCA 2251 TTGAGCAAGA CCGACATCAG AGGCAATGTCAGCCTTGCCG ATCACGCTCA 2301 TTTAAATCTC ACAGGACTTG CCACACTCAA CGGCAATCTTAGTGCAGGCG 2351 GAGACACGCA CTATACGGTT ACGCGCAACG CCACCCAAAA CGGCAACCTC2401 AGCCTCGTGG GCAATGCCCA AGCAACATTT AATCAAGCCA CATTAAACGG 2451CAACACATCG GCTTCGGACA ATGCTTCATT TAATCTAAGC AACAACGCCG 2501 TACAAAACGGCAGTCTGACG CTTTCCGACA ACGCTAAGGC AAACGTAAGC 2551 CATTCCGCAC TCAACGGCAATGTCTCCCTA GCCGATAAGG CAGTATTCCA 2601 TTTTGAAAAC AGCCGCTTTA CCGGAAAAATCAGCGGCGGC AAGGATACGG 2651 CATTACACTT AAAAGACAGC GAATGGACGC TGCCGTCGGGCACGGAATTA 2701 GGCAATTTAA ACCTTGACAA CGCCACCATT ACACTCAATT CCGCCTATCG2751 ACACGATGCG GCAGGCGCGC AAACCGGCAG TGCGGCAGAT GCGCCGCGCC 2801GCCGTTCGCG CCGTTCCCTA TTATCCGTTA CGCCGCCAAC TTCGGCAGAA 2851 TCCCGTTTCAACACGCTGAC GGTAAACGGC AAATTGAACG GTCAGGGAAC 2901 ATTCCGCTTT ATGTCGGAACTCTTCGGCTA CCGCAGCGGC AAATTGAAGC 2951 TGGCGGAAAG TTCCGAAGGC ACTTACACCTTGGCTGTCAA CAATACCGGC 3001 AACGAACCCG TAAGTCTCGA GCAATTGACG GTAGTGGAAGGAAAAGACAA 3051 CACACCGCTG TCCGAAAATC TTAATTTCAC CCTGCaaaAc gaacacgtcg3101 atgccggcgc atggCGTTAT CAGCTTATCC gcaaagacgG CGAGTTCCgc 3151CTGCATAATC CGGTCAAAGA ACAAGAGCTT TCCGACAAAC TCGGCAAGgc 3201 gggagaaACAGAggccgccT TGACGGCAAA ACAGGCacaA CTTGCCGCCA 3251 AAcaacaggc ggaaaAAGACAACgcgcaaa gccttgAcgc gctgattgcg 3301 gCcgggcgca atgccaccga AAAGGCAgaaagtgttgccg aaccgGCCCG 3351 GCAGGCAGGC GGGGAAAAtg ccgGCATTAT GCAGGCGGAGGAAGAGAAAA 3401 AACGGGTGCA GGCGGATAAA GACACCGCCT TGGCGAAACA GCGCGAAGCG3451 GAAACCCGGC CGGCTACCAC CGCCTTCCCC CGCGCCCGCC GCGCCCGCCG 3501GGATTTGCCG CAACCGCAGC CCCAACCGCA ACCCCAACCG CAGCGCGACC 3551 TGATCAGCCGTTATGCCAAT AGCGGTTTGA GTGAATTTTC CGCCACGCTC 3601 AACAGCGTTT TCGCCGTACAGGACGAATTG GACCGCGTGT TTGCCGAAGA 3651 CCGCCGCAAC GCCGTTTGGA CAAGCGGCATCCGGGACACC AAACACTACC 3701 GTTCGCAAGA TTTCCGCGCC TACCGCCAAC AAACCGACCTGCGCCAAATC 3751 GGTATGCAGA AAAACCTCGG CAGCGGGCGC GTCGGCATCC TGTTTTCGCA3801 CAACCGGACC GGAAACACCT TCGACGACGG CATCGGCAAC TCGGCACGGC 3851TTGCCCACGG TGCCGTTTTC GGGCAATACG GCATCGGCAG GTTCGACATC 3901 GGCATCAGCGCGGGCGCGGG TTTTAGTAGC GGCAGCCTTT CAGACGGCAT 3951 CAGAGGCAAA ATCCGCCGCCGCGTGCTGCA TTACGGCATT CAGGCAAGAT 4001 ACCGCGCAGG TTTCGGCGGA TTCGGCATCGAACCGCACAT CGGCGCAACG 4051 CGCTATTTCG TCCAAAAAGC GGATTACCGA TACGAAAACGTCAATATCGC 4101 CACCCCGGGC CTTGCATTCA ACCGCTACCG CGCGGGCATT AAGGCAGATT4151 ATTCATTCAA ACCGGCGCAA CACATTTCCA TCACGCCTTA TTTGAGCCTG 4201TCCTATACCG ATGCCGCTTC CGGCAAAGTC CGAACGCGCG TCAATACCGC 4251 CGTATTGGCGCAGGATTTCG GCAAAACCCG CAGTGCGGAA TGGGGCGTAA 4301 ACGCCGAAAT CAAAGGTTTCACGCTGTCCC TCCACGCTGC CGCCGCCAAG 4351 GGGCCGCAAT TGGAAGCGCA GCACAGCGCGGGCATCAAAT TAGGCTACCG 4401 CTGGTAA

This is predicted to encode a protein having amino acid sequence <SEQ ID654>:

1 MKTTDKRTTE THRKAPKTGR IRFSPAYLAI CLSFGILPQA RAGHTYFGIN 51 YQYYRDFAENKGKFAVGAKD IEVYNKKGEL VGKSMTKAPM IDFSVVSRNG 101 VAALAGDQYI VSVAHNGGYNNVDFGAEGSN PDQHRFSYQI VKRNNYKAGT 151 NGHPYGGDYH MPRLHKFVTD AEPVEMTSYMDGWKYADLNK YPDRVRIGAG 201 RQYWRSDEDE PNNRESSYHI ASAYSWLVGG NTFAQNGSGGGTVNLGSEKI 251 KHSPYGFLPT GGSFGDSGSP MFIYDAQKQK WLINGVLQTG NPYIGKSNGF301 QLVRKDWFYD EIFAGDTHSV FYEPHQNGKY FFNDNNNGAG KIDAKHKHYS 351LPYRLKTRTV QLFNVSLSET AREPVYHAAG GVNSYRPRLN NGENISFIDK 401 GKGELILTSNINQGAGGLYF EGNFTVSPKN NETWQGAGVH ISDGSTVTWK 451 VNGVANDRLS KIGKGTLLVQAKGENQGSVS VGDGKVILDQ QADDQGKKQA 501 FSEIGLVSGR GTVQLNADNQ FNPDKLYFGFRGGRLDLNGH SLSFHRIQNT 551 DEGAMIVNHN QDKESTVTIT GNKDITTTGN NNNLDSKKEIAYNGWFGEKD 601 ATKTNGGLNL NYPPEEADRT LLLSGGTNLN GNITQTNGKL FFSGRPTPHA651 YNHLGSGWSK MEGIPQGEIV WDNDWIDRTF KAENFHIQGG QAVVSRNVAK 701VEGDWHLSNH AQAVFGVAPH QSHTICTRSD WTGLTSCTEK TITDDKVIAS 751 LSKTDVRGNVSLADHAHLNL TGLATFNGNL VQAETRTIRL RANATQNGNL 801 SLVGNAQATF NQATLNGNTSASDNASFNLS NNAVQNGSLT LSDNAKANVS 851 HSALNGNVSL ADKAVFHFEN SRFTGKISGGKDTALHLKDS EWTLPSGTEL 901 GNLNLDNATI TLNSAYRHDA AGAQTGSAAD APRRRSRRSLLSVTPPTSAE 951 SRFNTLTVNG KLNGQGTFRF MSELFGYRSG KLKLAESSEG TYTLAVNNTG1001 NEPVSLEQLT VVEGKDNTPL SENLNFTLQN EHVDAGAWRY QLIRKDGEFR 1051LHNPVKEQEL SDKLGKAGET EAALTAKQAQ LAAKQQAEKD NAQSLDALIA 1101 AGRNATEKAESVAEPARQAG GENAGIMQAE EEKKRVQADK DTALAKQREA 1151 ETRPATTAFP RARRARRDLPQPQPQPQPQP QRDLISRYAN SGLSEFSATL 1201 NSVFAVQDEL DRVFAEDRRN AVWTSGIRDTKHYRSQDFRA YRQQTDLRQI 1251 GMQKNLGSGR VGILFSHNRT GNTFDDGIGN SARLAHGAVFGQYGIGRFDI 1301 GISAGAGFSS GSLSDGIRGK IRRRVLHYGI QARYRAGFGG FGIEPHIGAT1351 RYFVQKADYR YENVNIATPG LAFNRYRAGI KADYSFKPAQ HISITPYLSL 1401SYTDAASGKV RTRVNTAVLA QDFGKTRSAE WGVNAEIKGF TLSLHAAAAK 1451 GPQLEAQHSAGIKLGYRW*

Underlined and double-underlined sequences represent the active site ofa serine protease (trypsin family) and an ATP/GTP-binding site motif A(P-loop).

ORF1-1 and ORF1ng show 93.7% identity in 1471 aa overlap:

In addition, ORF1ng shows 55.7% identity with hap protein (P45387) overa 1455aa overlap:

Based on this analysis, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 78

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 655>:

1 ..AAGGTGTGGC AATTTGTCGA AGA.CCGCTG CGTGCCGTCG TGCCTGCCGA 51  CAGTTTTGAA CCGACCGCGC AAAAATTGAA CCTGTTTAAG GCGGGTGCGG 101  CAACCATTTT GTTTTATGAA GATCAAAATG TCGTCAAAGG TTTGCAGGAG 151  CAGTTCCCTG CTTATGCCGC TAACTTCCCC GTTTGGGCGg ATCAGGCAAA 201  CGCGATGGTG CAGTATGCCG TTTGGACGAC ACTTGCCGCG GTCGGCGTAG 251  GTGCAAACCT GCAACATTAC AATCCCTTGC CCGATGCGGC GATTGCCAAA 301  GCGTGGAATA TCCCCGAAAA CTGGTTGTTG CGCGCACAAA TGGTTATCGG 351  CGGTATTGAA GGGGCGGCAG GTGAAAAGAC CTTTGAACCC GTTGCAGAAC 401  GTTTGAAAGT GTTCGGCGCA TAA

This corresponds to the amino acid sequence <SEQ ID 656; ORF6>:

  1 . . . KVWQFVEXPL RAWPADSFE PTAQKLNLFK AGAATILFYE  DQNVVKGLQE  51      QFPAYAANFP VWADQANAMV QYAVWTTLAA VGVGANLQHY NPLPDAAIAK 101      AWNIPENWLL RAQMVIGGIE GAAGEKTFEP VAERLKVFGA *

Further sequence analysis revealed a further partial DNA sequence <SEQID 657>:

  1 . . . CTGCGTGCCG TCGTGCCTGC CGACAGTTTT GAACCGACCG CGCAAAAATT  51      GAACCTGTTT AAGGCGGGTG CGGCAACCAT TTTGTTTTAT GAAGATCAAA 101      ATGTCGTCAA AGGTTTGCAG GAGCAGTTCC CTGCTTATGC CGCTAACTTC 151      CCCGTTTGGG CGGATCAGGC AAACGCGATG GTGCAGTATG CCGTTTGGAC 201      GACACTTGCC GCGGTCGGCG TAGGTGCAAA CCTGCAACAT TACAATCCCT 251      TGCCCGATGC GGCGATTGCC AAAGCGTGGA ATATCCCCGA AAACTGGTTG 301      TTGCGCGCAC AAATGGTTAT CGGCGGTATT GAAGGGGCGG CAGGTGAAAA 351      GACCTTTGAA CCCGTTGCAG AACGTTTGAA AGTGTTCGGC GCATAA

This corresponds to the amino acid sequence <SEQ ID 658; ORF6-1>:

 1 . . . LRAVVPADSF EPTAQKLNLF KAGAATILFY EDQNVVKGLQ EQFPAYAANF  51      PVWADQANAM VQYAVWTTLA AVGVGANLQH YNPLPDAAIA KAWNIPENWL 101      LRAQMVIGGI EGAAGEKTFE PVAERLKVFG A*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF6 shows 98.6% identity over a 140aa overlap with an ORF (ORF6a) fromstrain A of N. meningitidis:

The complete length ORF6a nucleotide sequence <SEQ ID 659> is:

  1 ATGACCCGTC AATCTCTGCA ACAGGCTGCC GAAAGCCGCC GTTCCATTTA  51TTCGTTAAAT AAAAATCTGC CCGTCGGCAA AGATGAAATC GTCCAAATCG 101TCGAACACGC CGTTTTGCAC ACACCTTCTT CGTTCAATTC CCAATCTGCC 151CGTGTGGTCG TGCTGTTTGG CGAAGAGCAT GATAAGGTGT GGCAATTTGT 201CGAAGACGCG CTGCGTGCCG TCGTGCCTGC CGACAGTTTT GAACCGACCG 251CGCAAAAATT GAACCTGTTT AAGGCGGGTG CGGCAACTAT TTTGTTTTAT 301GAAGATCAAA ATGTCGTCAA AGGTTTGCAG GAGCAGTTCC CTGCTTATGC 351CGCCAACTTT CCCGTTTGGG CGGACCAGGC GAACGCGATG GTGCAGTATG 401CCGTTTGGAC GACACTTGCC GCGGTCGGCG TAGGTGCAAA CCTGCAACAT 451TACAATCCCT TGCCCGATGC GGCGATTGCC AAAGCGTGGA ATATCCCCGA 501AAACTGGTTG TTGCGCGCAC AAATGGTTAT CGGCGGTATT GAAGGGGCGG 551CAGGTGAAAA GACCTTTGAA CCAGTTGCAG AACGTTTGAA AGTGTTCGGC 601 GCATAA

This is predicted to encode a protein having amino acid sequence <SEQ ID660>:

  1 MTRQSLQQAA ESRRSIYSLN KNLPVGKDEI VQIVEHAVLH TPSSFNSQSA  51RVVVLFGEEH DKVWQFVEDA LRAVVPADSF EPTAQKLNLF KAGAATILFY 101EDQNVVKGLQ EQFPAYAANF PVWADQANAM VQYAVWTTLA AVGVGANLQH 151YNPLPDAAIA KAWNIPENWL LRAQMVIGGI EGAAGEKTFE PVAERLKVFG 201 A*

ORF6a and ORF6-1 show 100.0% identity in 131 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF6 shows 95.7% identity over a 140aa overlap with a predicted ORF(ORF6ng) from N. gonorrhoeae:

The complete length ORF6ng nucleotide sequence <SEQ ID 661> wasidentified as:

  1 ATGGCCGTTG CGTCAAATGT CAGCTTGGAT ATGTCCAATC CTACGGTGTT  51ACGCATGGGA TTACCCTTAT ATATTGCGTC CCTAAGAAGG GGCGCAATAT 101ATAAGGTGTG GCAATTTGTC GAAGACGCGC TGCGTGCCGT CGTGCCTGCC 151GACAGTTTTG AACCGACCGC GCAAAAATTG AAGCTGTTTA AGGCGGGCGC 201GGCAACCATT TTGTTTTATG AAGATCAAAA TGTCGTCAAA GGTTTGCAGG 251AGCAGTTCCC TGCTTATGCC GCCAACTTTC CCGTTTGGGC GGACCAGGCG 301AACGCTATGG TACAGTATGC CGTCTGGACG ACACTTGCCG CGGTCGGTGC 351AGGTGCAAAT CTGCAACATT ACAACCCCTT GCCCGATGTG GCGATTGCTA 401AAGCGTGGAA TATTCCCGAA AACTGGCTGT TGCGCGCGCA AATGGTTATC 451GGTGGTATTG AAGGGGcggc aggtgaaaaa gtctttgaac CCGTTGCgga 501acgtttgAAA GTGTTCGGCG CATAA

This encodes a protein having amino acid sequence <SEQ ID 662>:

  1 MAVASNVSLD MSNPTVLRMG LPLYIASLRR GAIYKVWQFV EDALRAVVPA  51DSFEPTAQKL KLFKAGAATI LFYEDQNVVK GLQEQFPAYA ANFPVWADQA 101NAMVQYAVWT TLAAVGAGAN LQHYNPLPDV AIAKAWNIPE NWLLRAQMVI 151GGIEGAAGEK VFEPVAERLK VFGA*

ORF6ng and ORF6-1 show 96.9% identity in 131 aa overlap:

It is predicted that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 79

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 663>

  1 . . . GGCTACAACT ACCTGTTCGC GCGCGGCAGC CGCATCGCCA ACTACCAAAT  51      CAACGGCATC CCCGTTGCCG ACGCGCTGGC CGATACGGGt CAATGCCAAC 101      ACCGCCGCCT ATGAGCGCGT AGAAGTCGTG CGCGGCGTGG CGGGGCTGCT 151      GGACGGCACG GGCGAGCCTT CCGCCACCGT CAATCTGGTG CGCAAACGCC 201      TGACCCGCAA GCCATTGTTT GAAGTCCGCG CCGAAGCgGG CAACCGcAAA 251      CATTTCGGGC TGGACGCGGA CGTATCGGGC AGCCTGAACA CCGAAG.crC 301      rCTGCGCgGC CGCCTGGTTT CCAcCTTCGG ACGCGGCGAC TCGTGGCGGC 351      GGCGCGAACG CAGCCGskAT GCCGAACTCT ACGGCATTTT GGAATACGAC 401      ATCGCACCGC AAACCCGCGT CCACGCArGC ATGGACTACC AGCAGGCGAA 451      AGAAACCGCC GACGCGCCGC TCAGcTACGC CGTGTACGAC AGCCAAGGTT 501      ATGCCACCGC CTTCGGCCCG AAAGACAACC CCGCCACAAA TTGGGCGAAC 551      AGCCACCACC GTGCGCTCAA CCTGTTCGCC GGCATCGAAC ACCGCTTCAA 601      CCAAGACTGG AAACTCAAAG CCGAATACGA CTAC . . . 

This corresponds to the amino acid sequence <SEQ ID 664; ORF23>:

  1 . . . GYNYLFARGS RIANYQINGI PVADALADTG NANTAAYERV EVVRGVAGLL  51      DGTGEPSATV NLVRKRLTRK PLFEVRAEAG NRKHFGLDAD VSGSLNTEXX 101      LRGRLVSTFG RGDSWRRRER SRXAELYGIL EYDIAPQTRV HAXMDYQQAK 151      ETADAPLSYA VYDSQGYATA FGPKDNPATN WANSHHRALN LFAGIEHRFN 201      QDWKLKAEYD Y . . . 

Further work revealed the complete nucleotide sequence <SEQ ID 665>:

   1 ATGACACGCT TCAAATATTC CCTGCTGTTT GCCGCCCTGT TGCCCGTGTA   51CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCCAAACCG CAGGAAAGCA  101CTGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC GAGTTCCAAC  151GACGGCTACA CTGTTTCCGG CACGCACACC CCGCTCGGGC TGCCCATGAC  201CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG CAACAAATGC  251GCGACCAAAA CATCAAAACG CTCGACCGCG CCCTGTTGCA GGCGACCGGC  301ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA ACTACCTGTT  351CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC ATCCCCGTTG  401CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC CTATGAGCGC  451GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CTGGACGGCA CGGGCGAGCC  501TTCCGCCACC GTCAATCTGG TGCGCAAACG CCTGACCCGC AAGCCATTGT  551TTGAAGTCCG CGCCGAAGCG GGCAACCGCA AACATTTCGG GCTGGACGCG  601GACGTATCGG GCAGCCTGAA CACCGAAGGC ACGCTGCGCG GCCGCCTGGT  651TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCGGCGCGAA CGCAGCCGCG  701ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC GCAAACCCGC  751GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG CCGACGCGCC  801GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC GCCTTCGGCC  851CGAAAGACAA CCCCGCCACA AATTGGGCGA ACAGCCGCCA CCGTGCGCTC  901AACCTGTTCG CCGGCATCGA ACACCGCTTC AACCAAGACT GGAAACTCAA  951AGCCGAATAC GACTACACCC GCAGCCGCTT CCGCCAGCCC TACGGCGTAG 1001CAGGCGTGCT TTCCATCGAC CACAACACCG CCGCCACCGA CCTGATTCCC 1051GGTTATTGGC ACGCCGACCC GCGCACCCAC AGCGCCAGCG TGTCATTGAT 1101CGGCAAATAC CGCCTGTTCG GCCGCGAACA CGATTTAATC GCGGGTATCA 1151ACGGTTACAA ATACGCCAGC AACAAATACG GCGAACGCAG CATCATCCCC 1201AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGTG CCTACCCGCA 1251GCCTGCATCG TTTGCCCAAA CCATCCCGCA ATACGGCACC AGGCGGCAAA 1301TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA CCTTTCGCTG 1351ATTTTGGGCG GACGATACAC CCGTTACCGC ACCGGCAGCT ACGACAGCCG 1401CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC CCCTACACAG 1451GCATCGTGTT CGACCTGACC GGCAACCTGT CTCTTTACGG CTCGTACAGC 1501AGCCTGTTCG TCCCGCAATC GCAAAAAGAC GAACACGGCA GCTACCTGAA 1551ACCCGTAACC GGCAACAATC TGGAAGCCGG CATCAAAGGC GAATGGCTTG 1601AAGGCCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG TAAAAACAAC 1651CTCGCCACCG CAGCAGGACG CGACCCGAGC GGCAACACCT ACTACCGCGC 1701CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC GGCGGCCGCA 1751TCACGCCCGA ATGGCAGATA CAGGCAGGTT ACAGCCAAAG CAAAACCCGC 1801GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTACCCG AACGCAGCTT 1851CAAACTCTTC ACTGCCTACC ACTTTGCCCC CGAAGCCCCC AGCGGCTGGA 1901CCATCGGCGC AGGCGTGCGC TGGCAGAGCG AAACCCACAC CGACCCTGCC 1951ACGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG CCGACAACAG 2001CCGCCAAAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT TACCGCTTCA 2051ATCCGCGCGC CGAACTGTCG CTGAACGTGG ACAATCTGTT CAACAAACAC 2101TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC GGACAGTGAA 2151CGCGGCGTTT ACCTATCGGT TTAAATAA

This corresponds to the amino acid sequence <SEQ ID 666; ORF23-1>:

  1 MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT VTADRTASSN  51DGYTVSGTHT PLGLPMTLRE IPQSVSVITS QQMRDQNIKT LDRALLQATG 101TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT GNANTAAYER 151VEVVRGVAGL LDGTGEPSAT VNLVRKRLTR KPLFEVRAEA GNRKHFGLDA 201DVSGSLNTEG TLRGRLVSTF GRGDSWRRRE RSRDAELYGI LEYDIAPQTR 251VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT NWANSRHRAL 301NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID HNTAATDLIP 351GYWHADPRTH SASVSLIGKY RLFGREHDLI AGINGYKYAS NKYGERSIIP 401NAIPNAYEFS RTGAYPQPAS FAQTIPQYGT RRQIGGYLAT RFRAADNLSL 451ILGGRYTRYR TGSYDSRTQG MTYVSANRFT PYTGIVFDLT GNLSLYGSYS 501SLFVPQSQKD EHGSYLKPVT GNNLEAGIKG EWLEGRLNAS AAVYRARKNN 551LATAAGRDPS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI QAGYSQSKTR 601DQDGSRLNPD SVPERSFKLF TAYHFAPEAP SGWTIGAGVR WQSETHTDPA 651TLRIPNPAAK ARAADNSRQK AYAVADIMAR YRFNPRAELS LNVDNLFNKH 701YRTQPDRHSY GALRTVNAAF TYRFK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Ferric-Pseudobactin Receptor PupB of Pseudomonasputida (Accession Number P38047)

ORF23 and PupB protein show 32% aa identity in 205aa overlap:

Orf23 6 FARGSRIANYQINGIPVADALADTGNANTAAYERVEVVRGVAGLLDGTGEPSATVNLVRK 65++RG  I NY+++G+P +   L D  + + A ++RVE+VRG  GL+ G G PSAT+NL+RK PupB 215WSRGFAIQNYEVDGVPTSTRL-DNYSQSMAMFDRVEIVRGATGLISGMGNPSATINLIRK 273 Orf2366 RLTRKPLFEVRAEAGNRKHFGLDADVSGSLNTEXXLRGRLVSTFXXXXXXXXXXXXXXAE 125R T +    +  EAGN   +G   DVSG L     +RGR V+ + PupB 274RPTAEAQASITGEAGNWDRYGTGFDVSGPLTETGNIRGRFVADYKTEKAWIDRYNQQSQL 333 Orf23126 LYGILEYDIAPQTRVHAXMDYQQAKETADAPLSYAVYD--SQGYATAFGPKDNPATNWAN 183+YGI E+D++  T+      Y   +    D+PL   +    S G  T      N A +W+ PupB 334MYGITEFDLSEDTLLTVGFSY--LRSDIDSPLRSGLPTRFSTGERTNLKRSLNAAPDWSY 391 Orf23184 SHHRALNLFAGIEHRFNQDWKLKAE 208 + H   + F  IE +    W  K E PupB 392NDHEQTSFFTSIEQQLGNGWSGKIE 416Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF23 shows 95.7% identity over a 211aa overlap with an ORF (ORF23a)from strain A of N. meningitidis:

The complete length ORF23a nucleotide sequence <SEQ ID 667> is:

   1 ATGACACGCT TCAAATATTC CCTGCTGTTT GCCGCCCTGT TGCCCGTGTA   51CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCAAAACCG CAGGAAAGCA  101CTGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC GAGTTCCAAC  151GACGGCTACA CTGTTTCCGG CACGCACACC CCGCTCGGGC TGCCCATGAC  201CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG CAACAAATGC  251GCGACCAAAA CATCAAAGCG CTCGACCGCG CCCTGTTGCA GGCGACCGGC  301ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA ACTACCTGTT  351CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC ATCCCCGTTG  401CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC CTATGAGCGC  451GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CTGGACGGCA CGGGCGAGCC  501TTCCGCCACC GTCAATCTGG TGCGCAAACG CCCGACCCGC AAGCCATTGT  551TTGAAGTCCG CGCCGAAGCG GGCAACCGCA AACATTTCGG GCTGGGCGCG  601GACGTATCGG GCAGCCTGAA TGCCGAAGGC ACGCTGCGCG GCCGCCTGGT  651TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCAGCGCGAA CGCAGCCGCG  701ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC GCAAACCCGC  751GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG CCGACGCGCC  801GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC GCCTTCGGCC  851CGAAAGACAA CCCCGCCACA AATTGGGCGA ACAGCCGCCA CCGTGCGCTC  901AACCTGTTCG CCGGCATCGA ACACCGCTTC AACCAAGACT GGAAACTCAA  951AGCCGAATAC GACTACACCC GCAGCCGCTT CCGCCAGCCC TACGGCGTAG 1001CAGGCGTGCT TTCCATCGAC CACAACACCG CCGCCACCGA CCTGATTCCC 1051GGTTATTGGC ACGCCGACCC GCGCACCCAC AGCGCCAGCG TGTCATTAAT 1101CGGCAAATAC CGCCTGTTCG GCCGCGAACA CGATTTAATC GCGGGTATCA 1151ACGGTTACAA ATACGCCAGC AACAAATACG GCGAACGCAG CATCATCCCC 1201AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGTG CCTACCCGCA 1251GCCTGCATCG TTTGCCCAAA CCATCCCGCA ATACGGCACC AGGCGGCAAA 1301TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA CCTTTCGCTG 1351ATACTCGGCG GCAGATACAG CCGTTACCGC ACCGGCAGCT ACGACAGCCG 1401CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC CCCTACACAG 1451GCATCGTGTT CGACCTGACC GGCAACCTGT CGCTTTACGG CTCGTACAGC 1501AGCCTGTTCG TCCCGCAATC GCAAAAAGAC GAACACGGCA GCTACCTGAA 1551ACCCGTAACC GGCAACAATC TGGAAGCCGG CATCAAAGGC GAATGGCTTG 1601AAGGCCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG TAAAAACAAC 1651CTCGCCACCG CAGCAGGACG CGACCCGAGC GGCAACACCT ACTACCGCGC 1701CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC GGCGGCCGCA 1751TCACGCCCGA ATGGCAGATA CAGGCAGGTT ACAGCCAAGG CAAAACCCGC 1801GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTACCCG AACGCAGCTT 1851CAAACTCTTC ACTGCCTACC ACTTTGCCCC CGAAGCCCCC AGCGGCTGGA 1901CCATCGGCGC AGGCGTGCGC TGGCAGAGCG AAACCCACAC CGACCCTGCC 1951ACGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG CCGACAACAG 2001CCGCCAAAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT TACCGCTTCA 2051ATCCGCGCGC CGAACTGTCG CTGAACGTGG ACAATCTGTT CAACAAACAC 2101TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC GGACAGTGAA 2151CGCGGCGTTT ACCTATCGGT TTAAATAA

This encodes a protein having amino acid sequence <SEQ ID 668>:

  1 MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT VTADRTASSN  51DGYTVSGTHT PLGLPMTLRE IPQSVSVITS QQMRDQNIKA LDRALLQATG 101TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT GNANTAAYER 151VEVVRGVAGL LDGTGEPSAT VNLVRKRPTR KPLFEVRAEA GNRKHFGLGA 201DVSGSLNAEG TLRGRLVSTF GRGDSWRQRE RSRDAELYGI LEYDIAPQTR 251VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT NWANSRHRAL 301NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID HNTAATDLIP 351GYWHADPRTH SASVSLIGKY RLFGREHDLI AGINGYKYAS NKYGERSIIP 401NAIPNAYEFS RTGAYPQPAS FAQTIPQYGT RRQIGGYLAT RFRAADNLSL 451ILGGRYSRYR TGSYDSRTQG MTYVSANRFT PYTGIVFDLT GNLSLYGSYS 501SLFVPQSQKD EHGSYLKPVT GNNLEAGIKG EWLEGRLNAS AAVYRARKNN 551LATAAGRDPS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI QAGYSQSKTR 601DQDGSRLNPD SVPERSFKLF TAYHFAPEAP SGWTIGAGVR WQSETHTDPA 651TLRIPNPAAK ARAADNSRQK AYAVADIMAR YRFNPRAELS LNVDNLFNKH 701YRTQPDRHSY GALRTVNAAF TYRFK*

ORF23a and ORF23-1 show 99.2% identity in 725 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF23 shows 93.4% identity over a 211 as overlap with a predicted ORF(ORF23.ng) from N. gonorrhoeae:

The ORF23ng nucleotide sequence <SEQ ID 669> is predicted to encode aprotein comprising amino acid sequence <SEQ ID 670>:

  1 SAVDACRIPG YNYLFARGSR IANYQINGIP VADALADTGN ANTAAYERVE  51VVRGVAGLPD GTGEPSATVN LVRKHPTRKP LFEVRAEAGN RKHFGLGADV 101SGSLNAEGTL RGRLVSTFGR GDSWRQLERS RDAELYGILE YDIAPQTRVH 151AGMDYQQAKE TADAPLSYAV YDSQGYATAF GPKDNPATNW SNSRNRALNL 201FAGIEHRFNQ DWKLKAEYDY TRSRFRQPYG VAGVLSIDHS TAATDLIPGY 251WHADPRTHSA SMSLTGKYRL FGREHDLIAG INGYKYASNK YGERSIIPNA 301IPNAYEFSRT GAYPQPSSFA QTIPQYDTRR QIGGYLATRF RAADNLSLIL 351GGRYSRYRAG SYNSRTQGMT YVSANRFTPY TGIVFDLTGN LSLYGSYSSL 401FVPQLQKDEH GSYLKPVTGN NLEADIKGEW LEGRLNASAA VYRARKNNLA 451TAAGRDQSGN TYYRAANQAK THGWEIEVGG RITPEWQIQA GYSQSKPRDQ 501DGSRLNPDSV PERSFKLFTA YHLAPEAPSG RTIGAGVRRQ GETHTDPAAL 551RIPNPAAKAR AVANSRQKAY AVADIMARYR FNPRTELSLN VDNLFNKHYR 601TQPDRHSYGA LRTVNAAFTY RFK*

Further work revealed the complete nucleotide sequence <SEQ ID 671>:

1 ATGACACGCT TCAAATACTC CCTGCTTTTT GCCGCCCTGC TACCCGTGTA 51CGCGCAGGCC GATGTTTCTG TTTCAGACGA CCCCAAACCG CAGGAAAGCA 101CCGAATTGCC GACCATCACC GTTACCGCCG ACCGCACCGC GAGTTCCAAC 151GACGGCTACA CCGTTTCCGG CACGCACACC CCGTTCGGGC TGCCCATGAC 201CCTGCGCGAA ATCCCGCAGA GCGTCAGCGT CATCACATCG CAACAAATGC 251GCGACCAAAA CATCAAAACG CTCGACCGCG CCCTGTTGCA GGCGACCGGC 301ACCAGCCGCC AGATTTACGG CTCCGACCGC GCGGGCTACA ACTACCTGTT 351CGCGCGCGGC AGCCGCATCG CCAACTACCA AATCAACGGC ATCCCCGTTG 401CCGACGCGCT GGCCGATACG GGCAATGCCA ACACCGCCGC CTATGAGCGC 451GTAGAAGTCG TGCGCGGCGT GGCGGGGCTG CCGGACGGCA CGGGCGAGCC 501TTCTGCCACC GTCAATCTGG TACGCAAACA CCCGACCCGC AAGCCATTGT 551TTGAAGTCCG CGCCGAAGCC GGCAACCGCA AACATTTCGG GCTGGGCGCG 601GACGTATCGG GCAGCCTGAA CGCCGAAGGC ACGCTGCGCG GCCGCCTGGT 651TTCCACCTTC GGACGCGGCG ACTCGTGGCG GCAGCTCGAA CGCAGCCGCG 701ATGCCGAACT CTACGGCATT TTGGAATACG ACATCGCACC GCAAACCCGC 751GTCCACGCAG GCATGGACTA CCAGCAGGCG AAAGAAACCG CAGACGCGCC 801GCTCAGCTAC GCCGTGTACG ACAGCCAAGG TTATGCCACC GCCTTCGGCC 851CAAAAGACAA CCCCGCCACA AATTGGTCGA ACAGCCGCAA CCGTGCGCTC 901AACCTGTTCG CCGGCATAGA ACACCGCTTC AACCAAGACT GGAAACTCAA 951AGCCGAATAC GACTACACCC GTAGCCGCTT CCGCCAGCCC TACGGTGTGG 1001CAGGCGTACT TTCCATCGAC CACAGCACTG CCGCCACCGA CCTGATTCCC 1051GGTTATTGGC ACGCcgatcc GCGCACCCAC AGCGCCAGCA TGTCATTGAC 1101CGGCAAATAC CgcctGTTCG GCCGCGAGCA CGATTTAATC GCGGGTATCA 1151ACGGCTACAA ATACGCCAGC AACAAATACG GCGAACGCAG CATCATTCCC 1201AACGCCATTC CCAACGCCTA CGAATTTTCC CGCACGGGCG CCTATCCGCA 1251GCCATCATCG TTTGCCCAAA CCATCCCGCA ATACGACACC AGGCGGCAAA 1301TCGGCGGCTA TCTCGCCACC CGTTTCCGCG CCGCCGACAA CCTTTCGCTG 1351ATACTCGGCG GCAGATACAG CCGCTACCGC GCAGGCAGCT ACAACAGCCG 1401CACACAAGGC ATGACCTATG TGTCCGCCAA CCGTTTCACC CCCTACACAG 1451GCATCGTGTT CGATCTGACC GGCAACCTGT CGCTTTACGG CTCGTACAGC 1501AGCCTGTTCG TCCCGCAATT GCAAAAAGAC GAACACGGCA GCTACCTGAA 1551ACCCGTAACC GGCAACAATC TGGAAGCCGA CATCAAAGGC GAATGGCTTG 1601AAGGGCGTCT GAACGCATCC GCCGCCGTGT ACCGCGCCCG TAAAAACAAC 1651CTCGCCACCG CAGCAGGACG CGACCAGAGC GGCAACACCT ACTATCGCGC 1701CGCCAACCAA GCCAAAACCC ACGGCTGGGA AATCGAAGTC GGCGGCCGCA 1751TCACGCCCGA ATGGCAGATA CAGGCAGGCT ACAGCCAAAG CAAACCCCGC 1801GACCAAGACG GCAGCCGCCT GAACCCCGAC AGCGTAcCCG AACGCAGCTT 1851CAAACTCTTC ACCGCCTACC ACTTAGCCCC CGAAGCCCCC AGCGGCCGGA 1901CCATcggTGC GGGTGTGCGC CGGCAGGGCG AAACCCACAC CGACCCAGCC 1951GCGCTCCGCA TCCCCAACCC CGCCGCCAAA GCCCGCGCCG TCGCCAACAG 2001CCGCCAGAAA GCCTACGCCG TCGCCGACAT CATGGCGCGT TACCGCTTCA 2051ATCCGCGCAC CGAACTGTCG CTGAACGTGG ACAACCTGTT CAACAAACAC 2101TACCGCACCC AGCCCGACCG CCACAGCTAC GGCGCACTGC GGACAGTGAA 2151CGCGGCGTTT ACCTATCGGT TTAAATAA

This corresponds to the amino acid sequence <SEQ ID 672; ORF23ng-1>:

  1 MTRFKYSLLF AALLPVYAQA DVSVSDDPKP QESTELPTIT VTADRTASSN  51DGYTVSGTHT PFGLPMTLRE IPQSVSVITS QQMRDQNIKT LDRALLQATG 101TSRQIYGSDR AGYNYLFARG SRIANYQING IPVADALADT GNANTAAYER 151VEVVRGVAGL PDGTGEPSAT VNLVRKHPTR KPLFEVRAEA GNRKHFGLGA 201DVSGSLNAEG TLRGRLVSTF GRGDSWRQLE RSRDAELYGI LEYDIAPQTR 251VHAGMDYQQA KETADAPLSY AVYDSQGYAT AFGPKDNPAT NWSNSRNRAL 301NLFAGIEHRF NQDWKLKAEY DYTRSRFRQP YGVAGVLSID HSTAATDLIP 351GYWHADPRTH SASMSLTGKY RLFGREHDLI AGINGYKYAS NKYGERSIIP 401NAIPNAYEFS RTGAYPQPSS FAQTIPQYDT RRQIGGYLAT RFRAADNLSL 451ILGGRYSRYR AGSYNSRTQG MTYVSANRFT PYTGIVFDLT GNLSLYGSYS 501SLFVPQLQKD EHGSYLKPVT GNNLEADIKG EWLEGRLNAS AAVYRARKNN 551LATAAGRDQS GNTYYRAANQ AKTHGWEIEV GGRITPEWQI QAGYSQSKPR 601DQDGSRLNPD SVPERSFKLF TAYHLAPEAP SGRTIGAGVR RQGETHTDPA 651ALRIPNPAAK ARAVANSRQK AYAVADIMAR YRFNPRTELS LNVDNLFNKH 701YRTQPDRHSY GALRTVNAAF TYRFK*

ORF23ng-1 and ORF23-1 show 95.9% identity in 725 aa overlap:

In addition, ORF23ng-1 shows significant homology with an OMP from E.coli:

sp|P16869|FHUE_ECOLI OUTER-MEMBRANE RECEPTOR FOR FE(III)-COPROGEN,FE(III)-FERRIOXAMINE B AND FE(III)-RHODOTRULIC ACID PRECURSOR >gi|1651542|gnl|PID|d1015403(D90745) Outer membrane protein FhuE precursor [Escherichia coli] >gi|1651545|gnl|PID|d1015405 (D90746) Outer membrane proteinFhuE precursor [Escherichia coli] >gi|1787344 (AE000210)outer-membrane receptor for Fe(III)-coprogen, Fe(III)-ferrioxamine B and Fe(III)-rhodotrulic acid precursor[Escherichia coli] Length = 729 Score = 332 bits (843), Expect = 3e−90Identities = 228/717 (31%), Positives = 350/717 (48%), Gaps =60/717 (8%) Query:  38TITVTADRTASSN--DGYTVSGTHTPFGLPMTLREIPQSVSVITSQQMRDQNIKTLDRAL  95 T+V    TA  +  + Y+V+ T     + MT R+IPQSV++++ Q+M DQ ++TL   + Sbjct:  43TVIVEGSATAPDDGENDYSVTSTSAGTKMQMTQRDIPQSVTIVSQQRMEDQQLQTLGEVM 102 Query: 96 LQATGTSRQIYGSDRAGYNYLFARGSRIANYQINGIP--------VADALADTGNANTAA 147    G S+    SDRA Y   ++RG +I NY ++GIP        + DAL+D      A Sbjct: 103ENTLGISKSQADSDRALY---YSRGFQIDNYMVDGIPTYFESRWNLGDALSDM-----AL 154 Query:148 YERVEVVRGVAGLPDGTGEPSATVNLVRKHPTRKPLF-EVRAEAGNRKHFGLGADVSGSL 206+ERVEVVRG  GL  GTG PSA +N+VRKH T +    +V AE G+       AD+   L Sbjct: 155FERVEVVRGATGLMTGTGNPSAAINMVRKHATSREFKGDVSAEYGSWNKERYVADLQSPL 214 Query:207 NAEGTLRGRLVSTFGRGDSWRQLERSRDAELYGILEYDIAPQTRVHAGMDYQQAKETADA 266  +G +R R+V  +   DSW     S      GI++ D+   T + AG +YQ+    + Sbjct: 215TEDGKIRARIVGGYQNNDSWLDRYNSEKTFFSGIVDADLGDLTTLSAGYEYQRIDVNSPT 274 Query:267 PLSYAVYDSQGYATAFGPKDNPATNWSNSRNRALNLFAGIEHRFNQDWKLKAEYDYTRSR 326      +++ G + ++    + A +W+ +      +F  ++ +F   W+      ++ Sbjct: 275WGGLPRWNTDGSSNSYDRARSTAPDWAYNDKEINKVFMTLKQQFADTWQATLNATHSEVE 334 Query:327 F--RQPYGVAGVLSIDHSTAA--TDLIPGY-------WHADPRTHSA-SMSLTGKYRLFG 374F  +  Y  A V   D       ++  PG+       W++  R   A  +   G Y LFG Sbjct: 335FDSKMMYVDAYVNKADGMLVGPYSNYGPGFDYVGGTGWNSGKRKVDALDLFADGSYELFG 394 Query:375 REHDLIAGINGYKYASNKYGER--SIIPNAIPNAYEFSRTGAYPQPSSFAQTIPQYDTRR 432R+H+L+ G   Y   +N+Y     +I P+ I + Y F+  G +PQ     Q++ Q DT Sbjct: 395RQHNLMFG-GSYSKQNNRYFSSWANIFPDEIGSFYNFN--GNFPQTDWSPQSLAQDDTTH 451 Query:433 QIGGYLATRFRAADNLSLILGGRYSRYRAGSYNSRTQGMTY-VSANRFTPYTGIVFDXXX 491    Y ATR   AD L LILG RY+ +R  +       +TY +  N  TPY G+VFD Sbjct: 452MKSLYAATRVTLADPLHLILGARYTNWRVDT-------LTYSMEKNHTTPYAGLVFDIND 504 Query:492 XXXXXXXXXXXFVPQLQKDEHGSYLKPVTGNNLEADIKGEWLEGRLNASAAVYRARKNNL 551           F PQ  +D  G YL P+TGNN E  +K +W+  RL  + A++R  ++N+ Sbjct: 505NWSTYASYTSIFQPQNDRDSSGKYLAPITGNNYELGLKSDWMNSRLTTTLAIFRIEQDNV 564 Query:552 ATAAGR---DQSGNTYYRAANQAKTHGWEIEVGGRITPEWQIQAGYSQSKPRDQDGSRLN 608 A +G      +G T Y+A +   + G E E+ G IT  WQ+  G ++    D +G+ +N Sbjct: 565AQSTGTPIPGSNGETAYKAVDGTVSKGVEFELNGAITDNWQLTFGATRYIAEDNEGNAVN 624 Query:609 PDSVPERSFKLFTAYHLAPEAPSGRTIGAGVRRQGETHTDPAALRIPNPAAKARAVANSR 668P ++P  + K+FT+Y L P  P   T+G GV  Q   +TD        P    RA Sbjct: 625P-NLPRTTVKMFTSYRL-PVMPE-LTVGGGVNWQNRVYTDTV-----TPYGTFRA----E 672 Query:669 QKAYAVADIMARYRFNPRTELSLNVDNLFNKHYRTQPDRH-SYGALRTVNAAFTYRF 724 Q +YA+D+  RY+      L  NV+NLF+K Y T  +    YG  R  +   TY+F Sbjct: 673QGSYALVDLFTRYQVTKNFSLQGNVNNLFDKTYDTNVEGSIVYGTPRNFSITGTYQF 729

Based on this analysis, it was predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF23-1 (77.5 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 15A shows the results ofaffinity purification of the His-fusion protein, and FIG. 15B shows theresults of expression of the GST-fusion in E. coli. Purified His-fusionprotein was used to immunise mice, whose sera were used for Western blot(FIG. 15C) and for ELISA (positive result). These experiments confirmthat ORF23-1 is a surface-exposed protein, and that it is a usefulimmunogen.

Example 80

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 673>:

  1 ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG CGGCTTCGTC  51GGCAATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG GGAACGGCAA 101TCATATCCAA GCCGACCGAA CAAACGGCGG TCATGGCTTC GAGTTTGTCC 151AGCGTCAgcA CGCCTGCTTC GGCGgcGgCa ATCATACCTT CGTCTTCGGA 201AACGGGGATA AACGcGCCAC TCAAACCCCC GACCGCGCTG GAAGCCATCA 251TGCCGCCTTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC TGCTGTTGTG 301CCGTGCGTAC CGCAGACGCT CAAGCCCATT TnTTCAAGAA TGCGTGCCAC 351TnAGTCGCCG ACGGGG..

This corresponds to the amino acid sequence <SEQ ID 674; ORF24>:

  1 MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISKPTE QTAVMASSLS  51SVSTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA SFSNAKAAVV 101PCVPQTLKPI XSRMRATXSP TG..

Further work revealed the complete nucleotide sequence <SEQ ID 675>:

  1 ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG CGGCTTCGTC  51GGCAATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG GGAACGGCAA 101TCATATCCAA GCCGACCGAA CAAACGGCGG TCATGGCTTC GAGTTTGTCC 151AGCGTCAGCA CGCCTGCTTC GGCGGCGGCA ATCATACCTT CGTCTTCGGA 201AACGGGGATA AACGCGCCAC TCAAACCCCC GACCGCGCTG GAAGCCATCA 251TGCCGCCTTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC TGCTGTTGTG 301CCGTGCGTAC CGCAGACGCT CAAGCCCATT TCTTCAAGAA TGCGTGCCAC 351TGAGTCGCCG ACGGCGGGGG TCGGCGCCAG CGACAAGTCG AGAATACCAA 401ACGGGATATT CAGCATTTTT GAGGCTTCGC GGCCGATGAG TTCGCCCACG 451CGGGTAATTT TGAAAGCAGT TTTCTTCACT ACTTCCGCAA CTTCGGTCAA 501TGTCGTTGCA TCTGAATTTT CCAACGCGGC TTTTACGACA CCTGGGCCGG 551ATACGCCGAC ATTGATAACG GCATCCGCTT CGCCCGAACC ATGAAACGCG 601CCCGCCATAA ACGGGTTGTC TTCCACCGCG TTGCAGAACA CGACAATTTT 651AGCGCAGCCG AAACCTTCGG GCGTGATTTC CGCCGTGCGT TTGACGGTTT 701CGCCCGCCAG CTTGACCGCA TCCATATTGA TACCGGCACG CGTACTGCCG 751ATATTGATGG AGCTGCACAC AATATCGGTA GTCTTCATCG CTTCGGGAAT 801GGAGCGGATT AACACCTCAT CCGAAGGCGA CATCCCTTTT TGCACCAACG 851CGGAAAAACC GCCGATAAAA GACACACCGA TGGCTTTGGC AGCTTTATCC 901AAAGTTTGCG CCACGCTGAC GTAA

This corresponds to the amino acid sequence <SEQ ID 676; ORF24-1>:

  1 MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISKPTE QTAVMASSLS  51SVSTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA SFSNAKAAVV 101PCVPQTLKPI SSRMRATESP TAGVGASDKS RIPNGIFSIF EASRPMSSPT 151RVILKAVFFT TSATSVNVVA SEFSNAAFTT PGPDTPTLIT ASASPEP*NA 201PAINGLSSTA LQNTTILAQP KPSGVISAVR LTVSPASLTA SILIPARVLP 251ILMELHTISV VFIASGMERI NTSSEGDIPF CTNAEKPPIK DTPMALAALS 301 KVCATLT*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF24 shows 96.4% identity over a 307 aa overlap with an ORF (ORF24a)from strain A of N. meningitidis:

The complete length ORF24a nucleotide sequence <SEQ ID 677> is:

  1 ATGCGCACGG CAGTGGTTTT GCTGTTGATC ATGCCGATGG CGGCTTCGTC  51GGCAATGATG CCGGAAATGG TGTGCGCGGG TGTGTCGCCG GGAACGGCAA 101TCATATCCAA NCCGACCGAA CAAACGGCGG TCATCGCTTC GAGTTTATCC 151AACGTCAGCA CGCCTGCTTC GGCGGCGGCA ATCATACCTT CGTCTTCGGA 201NACGGGGATA AACGCGCCAC TCAAACCGCC AACCGCGCTC GAAGCCATCA 251TGCCGCCCTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC TGCTGTTGTG 301CCGTGCGTAC CGCAGACGCT CAAACCCATT TCTTCAAGAA TGCGCGCCAC 351CGAGTCGCCG ACGGCAGGGG TCGGTGCCAG CGACAAGTCG AGAATACCAA 401ACGGGATATT CAGCATTTTT GAGGCTTCGC GGCCGATGAG TTCGCCCACG 451CGGGTAATTT TGAAGGCGGT TTTCTTCACA ACTTCGGCAA CTTCGGTCAA 501TGTCGTTGCA TCCGAATTTT CCAACGCGGC TTTTACGACA CCCGGGCCGG 551ATACGCCGAC ATTAATCACA GCATCCGCTT CGCCTGAGCC GTGAAACGCG 601CCCGCCATAN ACGGGTTGTC TTCCNCCGCG TTGCAGAACA CGACGATTTT 651GGCGCAGCCG AAACCTTCTA GTGTGATTTC ANCCGTGCGT TTGATGGTTT 701CGCCCGCCAG TCTGACCGCG TCCATATTGA TACCGGCGCG CGTACTGCCG 751ATATTGATGG AGCTGCACAC GATATCAGTA GTCTTCATCG CTTCGGGAAT 801GGAACGGATN AACACCTCGT CAGAAGGCGA CATACCTTTT TGCACCAGCG 851CGGAAAAGCC GCCAATAAAA GACACGCCGA TGGCTTTGGC AGCCTTATCC 901AAAGTTTGCG CCACGCTGAC GTAA

This encodes a protein having amino acid sequence <SEQ ID 678>:

  1 MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIISXPTE QTAVIASSLS  51NVSTPASAAA IIPSSSXTGI NAPLKPPTAL EAIMPPFFTA SFSNAKAAVV 101PCVPQTLKPI SSRMRATESP TAGVGASDKS RIPNGIFSIF EASRPMSSPT 151RVILKAVFFT TSATSVNVVA SEFSNAAFTT PGPDTPTLIT ASASPEP*NA 201PAIXGLSSXA LQNTTILAQP KPSSVISXVR LMVSPASLTA SILIPARVLP 251ILMELHTISV VFIASGMERX NTSSEGDIPF CTSAEKPPIK DTPMALAALS 301 KVCATLT*

It should be noted that this protein includes a stop codon at position198.

ORF24a and ORF24-1 show 96.4% identity in 307 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF24 shows 96.7% identity over a 121 aa overlap with a predicted ORF(ORF24ng) from N. gonorrhoeae:

The complete length ORF24ng nucleotide sequence <SEQ ID 679> is:

  1 ATGCGCACGG CGGTGGTTTT GCTGTTGATC ATGCCGATGG CGGCTTCGTC  51GGCGATGATG CCGGAAATGG TGTGCGCGGG CGTGTCGCCG GGAACGGCAA 101TCATGTCCAA ACCAACGGAG CAGACGGCGG TCATGGCTTC GAGTTTGTCC 151AGCGTCAACA CGCCTGCCTC GGCGGCGGCA ATCATACCTT CGTCTTCGGA 201AACGGGGATA AACGCGCCGC TCAAACCGCC GACCGCGCTG GAAGCCATCA 251TGCCGCCCTT TTTCACGGCA TCGTTCAGCA ATGCCAAAGC TGCTGTTGTG 301CCGTGCGTAC CGCAGACGCT CAAGCCCATT TCTTCAAGAA TGCGCGCCAC 351CGAGTCGCCG ACGGCGGGGG TCGGTGCCAG CGACAAATCG AGAATGCCGA 401ACGGGATATT CAGCATTTTT GAGGCTTCGC GACCGATGAG TTCGCCCACG 451CGGGTGATTT TGAAAGCGGT TTTCTTCACG ACTTCGGCGA CCTCGGTCAG 501GCTGACCGCG TCCGAATTTT CCAGCGCGGC TTTGACCACG CCTGGACCGG 551ATACGCCGAC ATTAATCACA GCATCCGCTT CGCCCGAGCC GTGGAACGCA 601CCCGCCATAA ACGGATTGTC TTCCACCGCG TTGCAGAACA CGACGATTTT 651GGCGCAGCCG AAACCTTCGG GTGTGATTTC AGCCGTGCGT TTGATGGTTT 701CGCCTGCCAG CTTGACCGCA TCCATATTGA TACCGGCACG CGTGCTGCCG 751ATATTGATGG AGCTGCACAC GATATCGGTA GTTTTCATCG CTTCGGGAAC 801GGAACGGATC AACACCTCAT CCGAAGGCGA CATACCTTTT TGCACCAGCG 851CGGAAAAGCC GCCGATAAAG GACACGCCGA TGGCTTTGGC TGCCTTGTCC 901AAAGTCTGCG CCACGCTGAC ATAA

This encodes a protein having amino acid sequence <SEQ ID 680>:

  1 MRTAVVLLLI MPMAASSAMM PEMVCAGVSP GTAIMSKPTE QTAVMASSLS  51SVNTPASAAA IIPSSSETGI NAPLKPPTAL EAIMPPFFTA SFSNAKAAVV 101PCVPQTLKPI SSRMRATESP TAGVGASDKS RMPNGIFSIF EASRPMSSPT 151RVILKAVFFT TSATSVRLTA SEFSSAALTT PGPDTPTLIT ASASPEPWNA 201PAINGLSSTA LQNTTILAQP KPSGVISAVR LMVSPASLTA SILIPARVLP 251ILMELHTISV VFIASGTERI NTSSEGDIPF CTSAEKPPIK DTPMALAALS 301 KVCATLT*

ORF24ng and ORF24-1 show 96.1% identity in 307 aa overlap:

Based on this analysis, including the presence of a putative leadersequence (first 18 aa—double-underlined) and putative transmembranedomains (single-underlined) in the gonococcal protein, it is predictedthat the proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 81

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 681>:

  1 ..ACCGACGTGC AAAAAGAGTT GGTCGGCGAA CAACGCAAGT GGGCGCAGGA  51  AAAAATCAGC AACTGCCGAC AAGCCGCCGC GCAGGCAGAC CGGCAGGAAT 101  ACGCCGAATA CCTCAAGCTG CAATGCGACA CGCGGATGAC GCGCGAACGG 151  ATACAGTATC TTCGCGGCTA TTCCATCGAT TAG

This corresponds to the amino acid sequence <SEQ ID 682; ORF25>:

-   -   1 . . . TDVQKELVGE QRKWAQEKIS NCRQAAAQAD RQEYAEYLKL QCDTRMTRER    -   51 IQYLRGYSID *

Further work revealed the complete nucleotide sequence <SEQ ID 683>:

  1 ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC TTGCCGCTTG  51CGGCAGGGAA GAACCGCCCA AGGCATTGGA ATGCGCCAAC CCCGCCGTGT 101TGCAAGGCAT ACGCGGCAAT ATTCAGGAAA CGCTCACGCA GGAAGCGCGT 151TCTTTCGCGC GCGAAGACGG CAGGCAGTTT GTCGATGCCG ACAAAATTAT 201CGCCGCCGCC TACGGTTTGG CGTTTTCTTT GGAACACGCT TCGGAAACGC 251AGGAAGGCGG GCGCACGTTC TGTATCGCCG ATTTGAACAT TACCGTGCCG 301TCTGAAACGC TTGCCGATGC CAAGGCAAAC AGCCCCCTGT TGTACGGGGA 351AACTGCTTTG TCGGATATTG TGCGGCAGAA GACGGGCGGC AATGTCGAGT 401TTAAAGACGG CGTATTGACG GCAGCCGTCC GCTTCCTGCC CGTCAAAGAC 451GGTCAGACGG CATTTGTCGA CAACACGGTC GGTATGGCGG CGCAAACGCT 501GTCTGCCGCG CTGCTGCCTT ACGGCGTGAA GAGCATCGTG ATGATAGACG 551GCAAGGCGGT GAAAAAAGAA GACGCGGTCA GGATTTTGAG CGGAAAAGCC 601CGTGAAGAAG AACCGTCCAA ACCCACGCCC GAAGACATTT TGGAACACAA 651TGCCGCCGGC GGCGATGCGG GCGTACCCCA AGCCGCAGAA GGCGCGCCCG 701AACCGGAAAT CCTGCATCCT GACGACGGCG AGCGTGCCGA TACCGTTACC 751GTATCACGGG GCGAAGTGGA AGAGGCGCGC GTACAAAACC AGCGTGCGGA 801ATCCGAAATT ACCAAACTTT GGGGAGGACT CGATACCGAC GTGCAAAAAG 851AGTTGGTCGG CGAACAACGC AAGTGGGCGC AGGAAAAAAT CAGCAACTGC 901CGACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCG AATACCTCAA 951GCTGCAATGC GACACGCGGA TGACGCGCGA ACGGATACAG TATCTTCGCG 1001 GCTATTCCAT CGATTAG

This corresponds to the amino acid sequence <SEQ ID 684; ORF25-1>:

  1 MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQGIRGN IQETLTQEAR  51SFAREDGRQF VDADKIIAAA YGLAFSLEHA SETQEGGRTF CIADLNITVP 101SETLADAKAN SPLLYGETAL SDIVRQKTGG NVEFKDGVLT AAVRFLPVKD 151GQTAFVDNTV GMAAQTLSAA LLPYGVKSIV MIDGKAVKKE DAVRILSGKA 201REEEPSKPTP EDILEHNAAG GDAGVPQAAE GAPEPEILHP DDGERADTVT 251VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEQR KWAQEKISNC 301RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF25 shows 98.3% identity over a 60aa overlap with an ORF (ORF25a) fromstrain A of N. meningitidis:

The complete length ORF25a nucleotide sequence <SEQ ID 685> is:

  1 ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC TTGCCGCTTG  51CGGCAGGGAA GAACCGCCCA AGGCATTGGA ATGCGCCAAC CCCGCCGTGT 101TGCAANGCAT ACGCNGCAAT ATTCAGGAAA CGCTCACGCA GGAAGCGCGT 151TCTTTCGCGC GCGAAGACNG CANGCAGTTT GTCGATGCCG ACNAAATTAT 201CGCCGCCGCC TANGNTNNGN NGNTNTCTTT GGAACACGCT TCGGAAACGC 251AGGAAGGCGG GCGCACGTTC TGTNTCGCCG ATTTGAACAT TACCGTGCCG 301TCTGAAACGC TTGCCGATGC CAAGGCAAAC AGCCCCCTGC TGTACGGGGA 351AACCGCTTTG TCGGATATTG TGCGGCAGAA GACGGGCGGC AATGTCGAGT 401TTAAAGACGG CGTATTGACG GCAGCCGTCC GCTTCCTACC CGTCAAAGAC 451GGTCAGANGG CATTTGTCGA CAACACGGTC GGTATGGCGG CGCAAACGCT 501GTCTGCCGCG TTGCTGCCTT ACGGCGTGAA GAGCATCGTG ATGATAGACG 551GCAAGGCGGT AAAAAAAGAA GACGCGGTCA GGATTNTGAG CNGANAAGCC 601CGTGAANAAG AACCGTCCAA ANCCNNGCCC GAAGACATTT TGGAACATAA 651TGCCGCCGGA GGGGATGCAG ACGTACCCCA AGCCGGAGAA GACGCGCCCG 701AACCGGAAAT CCTGCATCCT GACGACGGCG AGCGTGCCGA TACCGTTACC 751GTATCACGGG GCGAAGTGGA AGAGGCGCGN GTACAAAACC AGCGTGCGGA 801ATCCGAAATT ACCAAACTTT GGGGAGGACT CGATACCGAC GTGCAAAAAG 851AGTTGGTCGG CGAANAACGC AAGTGGGCGC AGGAAAAAAT CAGCAACTGC 901CGACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCG AATACCTCAA 951GCTGCAATGC GACACGCGGA TGACGCGCGA ACGGATACAG TATCTTCGCG 1001 GCTATTCCAT CGATTAG

This encodes a protein having amino acid sequence <SEQ ID 686>:

1 MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQXIRXN IQETLTQEAR 51 SFAREDXXQFVDADXIIAAA XXXXXSLEHA SETQEGGRTF CXADLNITVP 101 SETLADAKAN SPLLYGETALSDIVRQKTGG NVEFKDGVLT AAVRFLPVKD 151 GQXAFVDNTV GMAAQTLSAA LLPYGVKSIVMIDGKAVKKE DAVRIXSXXA 201 REXEPSKXXP EDILEHNAAG GDADVPQAGE DAPEPEILHPDDGERADTVT 251 VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEXR KWAQEKISNC301 RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

ORF25a and ORF25-1 show 93.5% identity in 338 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF25 shows 100% identity over a 60aa overlap with a predicted ORF(ORF25ng) from N. gonorrhoeae:

The complete length ORF25ng nucleotide sequence <SEQ ID 687> is:

1 ATGTATCGGA AACTCATTGC GCTGCCGTTT GCCCTGCTGC TTGCAGCGTG 51 CGGCAGGGAAGAACCGCCCA AGGCGTTGGA ATGCGCCAAC CCCGCCGTGT 101 TGCAGGACAT ACGCGGCAGTATTCAGGAAA CGCTCACGCA GGAAGCGCGT 151 TCTTTCGCGC GCGAAGACGG CAGGCAGTTTGTCGATGCCG ACAAAATTAT 201 CGCCGCCGCC TACGGTTTGG CGTTTTCTTT GGAACACGCTTCGGAAACGC 251 AGGAAGGCGG GCGCACGTTC TGTATCGCCG ATTTGAACAT TACCGTGCCG301 TCTGAAACGC TTGCCGATGC CGAGGCAAAC AGCCCCCTGC TGTATGGGGA 351AACGTCTTTG GCAGACATCG TGCAGCAGAA GACGGGCGGC AATGTCGAGT 401 TTAAAGACGGCGTATTGACG GCAGCCGTCC GCTTCCTGCC CGCCAAAGAC 451 GCTCGGACGG CATTTATCGACAACACGGTC GGTATGGCGA CGCAAACGCT 501 GTCTGCCGCG TTGCTGCCTT ACGGCGTGAAGAGCATCGTG ATGATAGACG 551 GCAAGGCGGT GACAAAAGAA GACGCGGTCA GGGTTTTGAGCGGCAAAGCC 601 CGTGAAGAAG AACCGTCCAA ACCCACCCCC GAAGACATTT TGGAACACAA651 TGCCGCCGGC GGCGATGCGG GCGTACCCCA AGCCGCAGAA GGCGCACCCG 701AACCCGAAAT CCTGCATCCC GACGACGTCG AGCGTGCCGA TACCGTTACC 751 GTATCACGGGGCGAAGTGGA AGAGGCGCGC GTACAAAACC AACGTGCGGA 801 ATCCGAAATT ACCAAACTTTGGGGAGGACT CGATACCGAC GTGCAAAAAG 851 AGTTGGTCGG CGAACAGCGC AAGTGGGCGCAGGAAAAAAT CAGcaactgc 901 cgACAAGCCG CCGCGCAGGC AGACCGGCAG GAATACGCCGAATACCTCAA 951 GCTCCAATGC GACACGCGGA TGACGCGCGA ACggaTACAG TATCTTCGCG1001 GCTATTCCAT CGATTAG

This encodes a protein having amino acid sequence <SEQ ID 688>:

1 MYRKLIALPF ALLLAACGRE EPPKALECAN PAVLQDIRGS IQETLTQEAR 51 SFAREDGRQFVDADKIIAAA YGLAFSLEHA SETQEGGRTF CIADLNITVP 101 SETLADAEAN SPLLYGETSLADIVQQKTGG NVEFKDGVLT AAVRFLPAKD 151 ARTAFIDNTV GMATQTLSAA LLPYGVKSIVMIDGKAVTKE DAVRVLSGKA 201 REEEPSKPTP EDILEHNAAG GDAGVPQAAE GAPEPEILHPDDVERADTVT 251 VSRGEVEEAR VQNQRAESEI TKLWGGLDTD VQKELVGEQR KWAQEKISNC301 RQAAAQADRQ EYAEYLKLQC DTRMTRERIQ YLRGYSID*

ORF25ng and ORF25-1 show 95.9% identity in 338 aa overlap:

Based on this analysis, including the presence of a predictedprokaryotic membrane lipoprotein lipid attchment site (underlined) inthe gonococcal protein, it was predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF25-1 (37 kDa) was cloned in pET and pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 16A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 16B shows theresults of expression of the His-fusion in E. coli. Purified His-fusionprotein was used to immunise mice, whose sera were used for Western blot(FIG. 16C), ELISA (positive result), and FACS analysis (FIG. 16D). Theseexperiments confirm that ORF25-1 is a surface-exposed protein, and thatit is a useful immunogen.

FIG. 16E shows plots of hydrophilicity, antigenic index, and AMPHIregions for ORF25-1.

Example 82

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 689>

1 ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG TGCCACCCTT 51 TTTGGCACTGGCACTTGCCG TCATTACCCG CCGCGTACTG CTGTCTTTAG 101 GCATCGGTAT TCTGGwysGCGTTGCCTTTT TGGTCGGCGG CAACCCCGTC 151 GACGGTCTGA CACACCTGAA AGACATGGTCGTCGGCTTGG CTTGGTCAGA 201 CGsyGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTCCkGATACTTT 251 TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA T.........                             // 851 .......... .......... ..................AC TTCGCTGGTA 901 TTCGGCGGCA CTTGCGGCGT CTTTGCCGTC GTTCTCTGCACGCTCGGCAC 951 GATTAAAACC GCCGACTATC CCAAAGCCGT TTGGCAGGGT GCGAAATCTA1001 TGTTCGGCGC AATCGCCATT TTAATCCTCG CTTGGCTCAT CAGTACGGTT 1051GTCGGCGAAA TGCACACCGG CGATTACCTC TCCACACTGG TTGCGGGCAA 1101 CATCCATCCCGGCTTCCTGC CCGTCATCCT CTTCCTGCTC GCCAGCGTGA 1151 TGGCGTTTGC CACAGGCACAAGCTGGGGGA CGTTCGGCAT TATGCTGCCG 1201 ATTGCCGCCG CCATGGCGGT CAAAGTCGAACCCGCGCTGA TTATCCCGTG 1251 TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCACTGCTCGCCCA 1301 TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG CAACCACATC1351 GACCACGTTA CCTCGCAACT GCCTTACGCC TTAACCGTTG CCGCCGCCGC 1401CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG CTGTTGGGCT 1451 TTGGCACGACAGGCATTGTA TTGGCGGTGC TGATTTTTCT GTTGAAAGAT 1501 AAAAAA..

This corresponds to the amino acid sequence <SEQ ID 690; ORF26>:

1 MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILXX VAFLVGGNPV 51 DGLTHLKDMVVGLAWSDXDW SLGKPKILVF XILLGIFTSL LTYSGSN...                             // 251 .......... .......... .................... ......TSLV 301 FGGTCGVFAV VLCTLGTIKT ADYPKAVWQG AKSMFGAIAILILAWLISTV 351 VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT SWGTFGIMLP401 IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL SSTGARCNHI 451DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV LAVLIFLLKD 501 KK..

Further work revealed the complete nucleotide sequence <SEQ ID 691>:

1 ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG TGCCACCCTT 51 TTTGGCACTGGCACTTGCCG TCATTACCCG CCGCGTACTG CTGTCTTTAG 101 GCATCGGTAT TCTGGTCGGCGTTGCCTTTT TGGTCGGCGG CAACCCCGTC 151 GACGGTCTGA CACACCTGAA AGACATGGTCGTCGGCTTGG CTTGGTCAGA 201 CGGCGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTCCTGATACTTT 251 TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA TCAGGCGTTT301 GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGCGCGGCG CGAAAATGCT 351GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT TTCCACAGTC 401 TCGCCGTCGGTGCGATTGCC CGCCCCGTTA CCGACAAGTT TAAAGTTTCC 451 CGCACCAAAC TCGCCTACATCCTCGACTCC ACTGCCGCTC CTATGTGCGT 501 GCTGATGCCC GTTTCAAGCT GGGGCGCGTCGATTATCGCC ACGCTTGCCG 551 GACTGCTCGT TACCTACAAA ATCACCGAAT ACACGCCGATGGGGACGTTT 601 GTCGCCATGA GCCTGATGAA CTATTACGCA CTGTTTGCCC TGATTATGGT651 GTTCGTCGTC GCATGGTTTT CCTTCGACAT CGGCTCGATG GCACGTTTCG 701AACAAGCCGC GTTGAACGAA GCCCACGATG AAACTGCCGT TTCAGACGCT 751 ACCAAAGGTCGTGTTTACGC ACTGATTATT CCCGTTTTGG CCTTAATCGC 801 CTCAACGGTT TCCGCCATGATCTACACCGG CGCGCAGGCA AGCGAAACCT 851 TCAGCATTTT GGGGGCATTT GAAAACACGGACGTAAACAC TTCGCTGGTA 901 TTCGGCGGCA CTTGCGGCGT CCTTGCCGTC GTTCTCTGCACGCTCGGCAC 951 GATTAAAACC GCCGACTATC CCAAAGCCGT TTGGCAGGGT GCGAAATCTA1001 TGTTCGGCGC AATCGCCATT TTAATCCTCG CTTGGCTCAT CAGTACGGTT 1051GTCGGCGAAA TGCACACCGG CGATTACCTC TCCACACTGG TTGCGGGCAA 1101 CATCCATCCCGGCTTCCTGC CCGTCATCCT CTTCCTGCTC GCCAGCGTGA 1151 TGGCGTTTGC CACAGGCACAAGCTGGGGGA CGTTCGGCAT TATGCTGCCG 1201 ATTGCCGCCG CCATGGCGGT CAAAGTCGAACCCGCGCTGA TTATCCCGTG 1251 TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCACTGCTCGCCCA 1301 TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG CAACCACATC1351 GACCACGTTA CCTCGCAACT GCCTTACGCC TTAACCGTTG CCGCCGCCGC 1401CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG CTGTTGGGCT 1451 TTGGCACGACAGGCATTGTA TTGGCGGTGC TGATTTTTCT GTTGAAAGAT 1501 AAAAAACGCG CCAACGCCTG A

This corresponds to the amino acid sequence <SEQ ID 692; ORF26-1>:

1 MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG VAFLVGGNPV 51 DGLTHLKDMVVGLAWSDGDW SLGKPKILVF LILLGIFTSL LTYSGSNQAF 101 ADWAKRHIKNRRGAKMLTAC LVFVTFIDDY FHSLAVGAIA RPVTDKFKVS 151 RTKLAYILDS TAAPMCVLMPVSSWGASIIA TLAGLLVTYK ITEYTPMGTF 201 VAMSLMNYYA LFALIMVFVV AWFSFDIGSMARFEQAALNE AHDETAVSDA 251 TKGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAFENTDVNTSLV 301 FGGTCGVLAV VLCTLGTIKT ADYPKAVWQG AKSMFGAIAI LILAWLISTV351 VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT SWGTFGIMLP 401IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL SSTGARCNHI 451DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV LAVLIFLLKD 501 KKRANA*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Hypothetical Transmembrane Protein HI1586 of H.influenzae (Accession Number P44263)

ORF26 and HI1586 show 53% and 49% amino acid identity in 97 and 221 aaoverlap at the N-terminus and C-terminus, respectively:

Orf26 1 MQLIDYSHSFFSVVPPFLALALAVITRRVXXXXXXXXXXXVAFLVGGNPVDGLTHLKDMV 60M+LID+S S +S+VP  LA+ LA+ TRRV              L          +L   V HI1586 14MELIDFSSSVWSIVPALLAIILAIATRRVLVSLSAGIIIGSLMLSDWQIGSAFNYLVKNV 73 Orf26 61VGLAWSDXDWSLGKPKILVFXILLGIFTSLLTYSGSN 97 V L ++D + +     I++F+LLG+ T+LLT SGSN HI1586 74 VSLVYADGEIN-SNMNIVLFLLLLGVLTALLTVSGSN 109                            // Orf26 86IFTSLLTYSGS--NTSLVFGGTCGVFAVVLCTL--GTIKTADYPKAVWQGAKSMFGXXXX 141 +F+ LT+  +   TSLV GG C +    L  +    +   +Y ++   G KSM G HI1586 299VFSVLGTFENTVVGTSLVVGGFCSIIISTLLIILDRQVSVPEYVRSWIVGIKSMSGAIAI 358 Orf26142 XXXXXXXSTVVGEMHTGDYLSTLVAGNIHPGFLPVILFLLASVMAFATGTSWGTFGIMLP 201       + +VG+M TG YLS+LV+GNI   FLPVILF+L + MAF+TGTSWGTFGIMLP HI1586 359LFFAWTINKIVGDMQTGKYLSSLVSGNIPMQFLPVILFVLGAAMAFSTGTSWGTFGIMLP 418 Orf26202 IAAAMAVKVEPALIIPCMSAVMAGAVCGDHCSPISDTTILSSTGARCNHIDHVTSQXXXX 261IAAAMA    P L++PC+SAVMAGAVCGDHCSP+SDTTILSSTGA+CNHIDHVT+Q HI1586 419IAAAMAANAAPELLLPCLSAVMAGAVCGDHCSPVSDTTILSSTGAKCNHIDHVTTQLPYA 478 Orf26262 XXXXXXXXXXXXXXXXXKSALLGFGTTGIVLAVLIFLLKDK 302                   S LGF  T + L V+IF +K + HI1586 479 ATVATATSIGYIVVGFTYSGLAGFAATAVSLIVIIFAVKKR519Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF26 shows 58.2% identity over a 502aa overlap with an ORF (ORF26a)from strain A of N. meningitidis:

The complete length ORF26a nucleotide sequence <SEQ ID 693> is:

1 ATGCAGCTGA TCGACTATTC ACATTCATTT TTCTCGGTTG TGCCACCCTT 51 TTTGGCACTGGCACTTGCCG TCATTACCCG CCGCGTACTG CTGTCTTTAG 101 GCATCGGTAT TCTGGTCGGCGTTGCCTTTT TGGTCGGCGG CAACCCCGTC 151 GACGGTCTGA CACACCTGAA AGACATGGTCGTCGGCTTGG CTTGGTCAGA 201 CGGCGATTGG TCGCTGGGCA AACCAAAANT CTTGGTTTTCCTGATACTTT 251 TGGGTATTTT TACTTCCCTG CTGACCTACT CCGGCAGCAA TCAGGCGTTT301 GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGCGCGGCG CGAAAATGCT 351GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT TTCCACAGTC 401 TCGCCGTCGGTGCGNTTGCC CGCCCCGTTA CCGACAAGTT TAAAGTTTCC 451 CGCGCCAAAC TCGCCTACATCCTCGACTCC ACTGCCGCGC CTATGTGCGT 501 GCTGATGCCC GTTTCAAGCT GGGGCGCGTCGATTATCGCC ACGCTTGCCG 551 GACTGCTCGT TACCTACAAA ATCACCGAAT ACACGCCGATGGGGACGTTT 601 GTCGCCATGA GCCTGATGAA CTATTACGCA CTGTTTGCCC TGATTATGGT651 GTTCGTCGTC GCATGGTTCT CCTTCGACAT CGGCTCGATG GCACGTTTCG 701AACAAGCCGC GTTGAACGAA GCCCACGATG AAACTGCCGT TTCAGACGGC 751 AGCTGGGGCAGGGTTTACGC ATTGATTATT CCCGTTTTGG CCTTAATCGC 801 CTCAACGGTT TCCGCCATGATCTACACCGG TGCACAGGCA AGCGAAACCT 851 TCAGCATTTT GGGTGCATTT GAAAATACGGACGTGAACAC TTCGCTGGTA 901 TTCGGCGGCA CTTGCGGCGT GCTTGCCGTC GTCCTCTGCACGCTCGGCAC 951 GATTAAAATC GCCGATTATC CCAAAGCCGT TTGGCAGGGT GCGAAATCCA1001 TGTTCGGCGC AATCGCCATT TTAATCCTTG CCTGGCTCAT CAGTACGGTT 1051GTCGGCGAAA TGCACACAGG CGACTACCTC TCCACGCTGG TTGCGGGCAA 1101 CATCCATCCCGGCTTCCTGN CCGTCATCCT TTTCCTGCTC GCCAGCGTGA 1151 TGGCGTTTGC CACAGGCACAAGCTGGGGGA CGTTCGGCAT CATGCTGCCG 1201 ATTGCCGCCG CCATGGCGGT CAAAGTCGATCCCTCACTGA TTATCCCGTG 1251 TATGTCCGCC GTGATGGCGG GGGCGGTATG CGGCGACCACTGCTCGCCCA 1301 TTTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG CAACCACATC1351 GACCACGTTA CNTCGCAACT GCCTTACGCC TTAACCGTTG CCGCCGCCGC 1401CGCATCGGGN TACCTCGCAT TGGGTCTGAC AAAATCCGCG CTGTTGGGTT 1451 TTGGCANGACAGGCATTGTA TTGGCGGTGC TGATTTTTCT GTTGAAAGAT 1501 AAAAAACGCG CCAACGCCTG A

This encodes a protein having amino acid sequence <SEQ ID 694>:

1 MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG VAFLVGGNPV 51 DGLTHLKDMVVGLAWSDGDW SLGKPKXLVF LILLGIFTSL LTYSGSNQAF 101 ADWAKRHIKNRRGAKMLTAC LVFVTFIDDY FHSLAVGAXA RPVTDKFKVS 151 RAKLAYILDS TAAPMCVLMPVSSWGASIIA TLAGLLVTYK ITEYTPMGTF 201 VAMSLMNYYA LFALIMVFVV AWFSFDIGSMARFEQAALNE AHDETAVSDG 251 SWGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAFENTDVNTSLV 301 FGGTCGVLAV VLCTLGTIKI ADYPKAVWQG AKSMFGAIAI LILAWLISTV351 VGEMHTGDYL STLVAGNIHP GFLXVILFLL ASVMAFATGT SWGTFGIMLP 401IAAAMAVKVD PSLIIPCMSA VMAGAVCGDH CSPISDTTIL SSTGARCNHI 451DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGXTGIV LAVLIFLLKD 501 KKRANA*

ORF26a and ORF26-1 show 97.8% identity in 506 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF26 shows 94.8% and 99% identity in 97 and 206 aa overlap at theN-terminus and C-terminus, respectively, with a predicted ORF (ORF26ng)from N. gonorrhoeae:

The complete length ORF26ng nucleotide sequence <SEQ ID 695> is:

1 ATGCAGCTGA TTGACTATTC ACATTCATTT TTCTCGGTTG TGCCACCCTT 51 TTTGGCACTGGCACTTGCCG TCATTACCCG CCGCGTACTG CTGTCTTTAG 101 GCATCGGTAT TTTGGTCGGCGTTGCCTTTT TGGTCGGCGG CAACCCCGTC 151 GACGGTCTGA CACACCTGAA AGACATGGTCGTCGGCTTGG CTTGGGCAGA 201 CGGCGATTGG TCGCTGGGCA AACCAAAAAT CTTGGTTTTCCTGATACTTT 251 TGGGCATTTT CACTTCACTG CTGACCTACT CCGGCAGCAA TCAGGCGTTT301 GCCGACTGGG CAAAACGGCA CATTAAAAAC CGGTGCGGCG CGAAAATGCT 351GACCGCCTGC CTCGTGTTCG TAACCTTTAT CGACGACTAT TTCCACAGCC 401 TCGCCGTCGGTGCGATTGCC CGCCCCGTTA CCGACAAGTT TAAAGTTTCC 451 CGCGCCAAAC TCGCCTACATCCTCGACTCC ACTGCCTCGC CCATGTGCGT 501 GCTGATGCCC GTTTCAAGCT GGGGCGCGTCGATTATCGCC ACGCTTGCCG 551 GATTGCTCGT TACCTACAAA ATTACCGAAT ACACGCCGATGGGGACGTTT 601 GTCGCCATGA GCCTGATGAA CTATTACGCG CTGTTTGCCC TGATTATGGT651 ATTCGTCGTC GCATGGTTCT CCTTCGACAT CGGCTCGAtg gCGCGTTTCG 701AACAGGCTGC GTTGAACGAA gcccaggacg aaaccgccgc tTCAGACgCT 751 ACCAAAGGTCGTGTTTACGC ATTGATTATT CCCGTTTTGG CCTTAATCGC 801 CTCAACGGTT TCCGCCATGATCTACACCGG CGCGCAGGCA AGCGAAACCT 851 TCAGCATTTT GGGGGCATTT GAAAATACCGACGTAAACAC TTCGCTGGTA 901 TTCGGCGGCA CTTGCGGCGT GCTTGCCGTC GTCCTCTGCACGTTCGGCAC 951 GATTAAAACC GCCGATTATC CCAAAGCCGT GTGGCAGGGT GCGAAATCCA1001 TGTTCGGCGC AATCGCCATT TTAATCCTCG CCTGGCTCAT CAGTACGGTT 1051GTCGGCGAAA TGCACACGGG CGACTACCTC TCCACGCTGG TTGCGGGCAA 1101 CATCCATCCCGGCTTCCTGC CCGTCATCCT CTTCCTGCTC GCCAGCGTGA 1151 TGGCGTTTGC CACAGGCACAAGCTGGGGGA CGTTCGGCAT TATGCTGCCG 1201 ATTGCCGCCG CCATGGCGGT CAAAGTCGAACCCGCGCTGA TTAtcccGTG 1251 TATGTCCGCA GTAATGGCGG GGGCGGTATG CGGCGACCACTGTTCGCCCA 1301 TCTCCGACAC GACCATCCTG TCGTCCACCG GCGCGCGCTG CAACCACATC1351 GACCACGTTA CCTCGCAACT GCCTTATGCC CTGACGGTTG CCGCCGCCGC 1401CGCATCGGGC TACCTCGCAT TGGGTCTGAC AAAATCCGCG CTGTTGGGCT 1451 TTGGCACGACCGGTATTGTA TTGGCGGTGC TGATTTTTCT GTTGAAAGAT 1501 AAAAAACGCG CCGACGTTTG A

This encodes a protein having amino acid sequence <SEQ ID 696>:

1 MQLIDYSHSF FSVVPPFLAL ALAVITRRVL LSLGIGILVG VAFLVGGNPV 51 DGLTHLKDMVVGLAWADGDW SLGKPKILVF LILLGIFTSL LTYSGSNQAF 101 ADWAKRHIKNRCGAKMLTAC LVFVTFIDDY FHSLAVGAIA RPVTDKFKVS 151 RAKLAYILDS TASPMCVLMPVSSWGASIIA TLAGLLVTYK ITEYTPMGTF 201 VAMSLMNYYA LFALIMVFVV AWFSFDIGSMARFEQAALNE AQDETAASDA 251 TKGRVYALII PVLALIASTV SAMIYTGAQA SETFSILGAFENTDVNTSLV 301 FGGTCGVLAV VLCTFGTIKT ADYPKAVWQG AKSMFGAIAI LILAWLISTV351 VGEMHTGDYL STLVAGNIHP GFLPVILFLL ASVMAFATGT SWGTFGIMLP 401IAAAMAVKVE PALIIPCMSA VMAGAVCGDH CSPISDTTIL SSTGARCNHI 451DHVTSQLPYA LTVAAAAASG YLALGLTKSA LLGFGTTGIV LAVLIFLLKD 501 KKRADV*

ORF26ng and ORF26-1 show 98.4% identity in 505 aa overlap:

In addition, ORF26 ng shows significant homology to a hypothetical H.influenzae protein:

sp|P44263|YF86_HAEIN HYPOTHETICAL PROTEIN HI1586 >gi|1074850|pir||C64037hypothetical protein HI1586 - Haemophilus influenzae (strain RdKW20) >gi|1574427 (U32832) H. influenzae predicted coding region HI1586[Haemophilus influenzae] Length = 519 Score = 538 bits (1370), Expect= e−152 Identities = 280/507 (55%), Positives = 346/507 (68%), Gaps= 7/507 (1%) Query: 1MQLIDYSHSFFSVVPPFLALALAVITRRXXXXXXXXXXXXXAFLVGGNPVDGLTHLKDMV 60 M+LID+SS +S+VP  LA+ LA+ TRR               L          +L   V Sbjct: 14MELIDFSSSVWSIVPALLAIILAIATRRVLVSLSAGIIIGSLMLSDWQIGSAFNYLVKNV 73 Query:61 VGLAWADGDWSLGKPKILVFLILLGIFTSLLTYSGSNQAFADWAKRHIKNRCGAKMLTAC 120 V L+ADG+ +     I++FL+LLG+ T+LLT SGSN+AFA+WA+  IK R GAK+L A Sbjct: 74VSLVYADGEIN-SNMNIVLFLLLLGVLTALLTVSGSNRAFAEWAQSRIKGRRGAKLLAAS 132 Query:121 LVFVTFIDDYFHSLAVGAIARPVTDKFKVSRAKLAYILDSTASPMCVLMPVSSWGASIIA 180LVFVTFIDDYFHSLAVGAIARPVTD+FKVSRAKLAYILDSTA+PMCV+MPVSSWGA II Sbjct: 133LVFVTFIDDYFHSLAVGAIARPVTDRFKVSRAKLAYILDSTAAPMCVMMPVSSWGAYIIT 192 Query:181 TLAGLLVTYKITEYTPMGTFVAMSLMNYYALFALIMVFVVAWFSFDIGSMARFEQAALNE 240 + GLL TY ITEYTP+G FVAMS MN+YA+F++IMVF VA+FSFDI SM R E+ AL Sbjct: 193LIGGLLATYSITEYTPIGAFVAMSSMNFYAIFSIIMVFFVAYFSFDIASMVRHEKLALKN 252 Query:241 AQDETAASDATKGRVYALIIPVLALIASTVSAMIYTGAQA----SETFSILGAFENTDVN 296 +D+      TKG+V  LI+P+L LI +TVS MIYTGA+A     + FS+LG FENT V Sbjct: 253TEDQLEEETGTKGQVRNLILPILVLIIATVSMMIYTGAEALAADGKVFSVLGTFENTVVG 312 Query:297 TSLVFGGTCGVL--AVVLCTFGTIKTADYPKAVWQGAKSMFGXXXXXXXXXXXSTVVGEM 354TSLV GG C ++   +++     +   +Y ++   G KSM G           + +VG+M Sbjct: 313TSLVVGGFCSIIISTLLIILDRQVSVPEYVRSWIVGIKSMSGAIAILFFAWTINKIVGDM 372 Query:355 HTGDYLSTLVAGNIHPGFLPVILFLLASVMAFATGTSWGTFGIMLPIAAAMAVKVEPALI 414  TGYLS+LV+GNI   FLPVILF+L + MAF+TGTSWGTFGIMLPIAAAMA    P L+ Sbjct: 373QTGKYLSSLVSGNIPMQFLPVILFVLGAAMAFSTGTSWGTFGIMLPIAAAMAANAAPELL 432 Query:415 IPCMSAVMAGAVCGDHCSPISDTTILSSTGARCNHIDHVTSQXXXXXXXXXXXXXXXXXX 474+PC+SAVMAGAVCGDHCSP+SDTTILSSTGA+CNHIDHVT+Q Sbjct: 433LPCLSAVMAGAVCGDHCSPVSDTTILSSTGAKCNHIDHVTTQLPYAATVATATSIGYIVV 492 Query:475 XXXKSALLGFGTTGIVLAVLIFLLKDK 501     S L GF  T + L V+IF +K + Sbjct:493 GFTYSGLAGFAATAVSLIVIIFAVKKR 519

Based on this analysis, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 83

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 697>:

1 ..AAGCAATGGT ATGCCGACGN .AGTATCAAG ACGGAAATGG TTATGGTCAA 51  CGATGAGCCT GCCAAAATTC TGACTTGGGA TGAAAGCGGC CGATTACTCT 101  CGGAACTGTC TATCCGCCAC CATCAACGCA ACGGGGTGGT TTTGGAGTGG 151  TATGAAGATG GTTCTAAAAA GAGCGAAGT. GTTTATCAGG ATGACAAGTT 201  GGTCAGGAAA ACCCAGTGGG ATAAGGATGG TTATTTAATC GAACCCTGA

This corresponds to the amino acid sequence <SEQ ID 698; ORF27>:

1 ..KQWYADXSIK TEMVMVNDEP AKILTWDESG RLLSELSIRH HQRNGVVLEW 51  YEDGSKKSEX VYQDDKLVRK TQWDKDGYLI EP*

Further work revealed the complete nucleotide sequence <SEQ ID 699>:

1 ATGAAAAAAT TATCTCGGAT TGTATTTTCA ACTGTCCTGT TGGGTTTTTC 51 GGCCGCTTTGCCGGCGCAGA CCTATTCTGT TTATTTTAAT CAGAACGGAA 101 AGCTGACGGC GACGATGTCTTCTGCCGCTT ATATCAGGCA ATATAGTGTG 151 GTGGCGGGTA TTGCGCACGC GCAGGATTTTTATTATCCGT CGATGAAGAA 201 ATATTCTGAA CCTTATATCG TTGCTTCAAC GCAAATCAAATCTTTTGTGC 251 CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA TGGTCAGAAA301 AAAATGGCGG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG AGTGGGTCAA 351CTGGTATCCG AACGGTAAAA AATCTGCCGT TATGCCTTAT AAAAATGGCT 401 TGAGTGAGGGTACGGGATAC CGCTATTACC GTAACGGCGG CAAGGAAAGC 451 GAAATCCAGT TTAAGCAAAATAAGGCAAAC GGCGTATGGA AGCAATGGTA 501 TGCCGACGGC AGTATCAAGA CGGAAATGGTTATGGTCAAC GATGAGCCTG 551 CCAAAATTCT GACTTGGGAT GAAAGCGGCC GATTACTCTCGGAACTGTCT 601 ATCCGCCACC ATCAACGCAA CGGGGTGGTT TTGGAGTGGT ATGAAGATGG651 TTCTAAAAAG AGCGAAGCTG TTTATCAGGA TGACAAGTTG GTCAGGAAAA 701CCCAGTGGGA TAAGGATGGT TATTTAATCG AACCCTGA

This corresponds to the amino acid sequence <SEQ ID 700; ORF27-1>:

1 MKKLSRIVFS TVLLGFSAAL PAQTYSVYFN QNGKLTATMS SAAYIRQYSV 51 VAGIAHAQDFYYPSMKKYSE PYIVASTQIK SFVPTLQNGM LILWHFNGQK 101 KMAGGFSKGK PDGEWVNWYPNGKKSAVMPY KNGLSEGTGY RYYRNGGKES 151 EIQFKQNKAN GVWKQWYADG SIKTEMVMVNDEPAKILTWD ESGRLLSELS 201 IRHHQRNGVV LEWYEDGSKK SEAVYQDDKL VRKTQWDKDGYLIEP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF27 shows 91.5% identity over a 82aa overlap with an ORF (ORF27a) fromstrain A of N. meningitidis:

The complete length ORF27a nucleotide sequence <SEQ ID 701> is:

1 ATGAAAAAAT TATCTCGGAT TGTATTTTCA ACTGTCCTGT TGGGTTTTTC 51 GGCCGCTTTGCCGGCGCAGA NCTATTCTGT TTATTTTAAT CAGAACGGGA 101 AACTGACGGC GACGNTGTCTTCTGCCGCNT ATATCAGGCA ATATAGTGTG 151 GCGGAGGGTA TTGCGCACGC GCAGGANTTTTANTATCCGT CGATGAAGAA 201 ATATTCCGAA CCTTATATCG TTGCTTCAAC GCAAATCAAATCTTTTGTGC 251 CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA NGGTCAGAAA301 AAAATGGCNG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG AGTGGGTCAA 351CTGGTATCCG AACGGTAAAA AATCTGCCGT TATGCCTTAT AAAAATGGTT 401 TGAGTGAAGGTACGGGGTNN CGCTATTACC GTAACGGCGG CAAGGAAAGC 451 GAAATCCAGT TTAAACAGAATAAGGCAAAC GGCGTATGGA AGCAATGGTA 501 TGCCGACGGC AATATCAAAA CGGAAATGGTTATGGTCAAT GATGAGCCTG 551 CCAAAATTCT GACATGGGAT GAAAGCGGTC GATTACTCTCGGAACTGTCT 601 ATCCATCATC ATNAACGTAA TGGAGTAGTC TTAGAGTGGT ATGAAGATGG651 TTCTAAAAAG ANTGAAGCTG TTTATCAGGA TGATAAGTTG GTCAGGAAAA 701CCCAGTGGGA TAANGATGGT TATTTAATCG AACCCTGA

This encodes a protein having amino acid sequence <SEQ ID 702>:

1 MKKLSRIVFS TVLLGFSAAL PAQXYSVYFN QNGKLTATXS SAAYIRQYSV 51 AEGIAHAQXFXYPSMKKYSE PYIVASTQIK SFVPTLQNGM LILWHFXGQK 101 KMAGGFSKGK PDGEWVNWYPNGKKSAVMPY KNGLSEGTGX RYYRNGGKES 151 EIQFKQNKAN GVWKQWYADG NIKTEMVMVNDEPAKILTWD ESGRLLSELS 201 IHHHXRNGVV LEWYEDGSKK XEAVYQDDKL VRKTQWDXDGYLIEP*

ORF27a and ORF27-1 show 94.7% identity in 245 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF27 shows 96.3% identity over 82 aa overlap with a predicted ORF(ORF27ng) from N. gonorrhoeae:

The complete length ORF27ng nucleotide sequence <SEQ ID 703> is:

1 ATGAAGAAAT TATCTCGGAT TGTATTTTCA ATCGTACTGT TGGGTTTTTC 51 GGCCGCTTTGCCGGCGCAGA CCTATTCTGT TTATTTTAAT CAGAACGGGA 101 AACTGACGGC GACGATGTCTTCTGCCGCTT ATATCAGGCA ATATAGTGTG 151 GCGGCGGGTA TCGCACACGC GCAGGATTTTTATTATCCGT CGATGAAGAA 201 ATATTCCGAA CCTTATATCG TTGCTTCAAC GCAAATCAAATCTTTTGTGC 251 CTACCCTGCA AAACGGTATG TTGATTTTGT GGCATTTTAA TGGTCAGAAA301 AAAATGGCGG GGGGCTTCAG CAAGGGTAAG CCGGACGGGG AATGGGTCAA 351CTGGTATCCG AACGGTAAAA AATCTGCGGT TATGCCTTAT AAAAATGGCT 401 TGAGTGAGGGTACGGGATAC CGTTATTACC GTAACGGCGG CAAGGAAAGC 451 GAAATCCAGT TTAAGCAAAATAAGGCGAAC GGCGTATGGA AGCAATGGTA 501 TGCCGATGGA AGTATCAAGA CGGAAATGGTTATGGTCAAC GATGAGCCTG 551 CCAAAATTCT GACTTGGGAT GAAAGCGGCC GATTACTTTCGGAACTGTCT 601 ATCCGCCACC ATAAACGCAA CGGGGTGGTT TTGGAGTGGT ATGAAGATGG651 TTCTAAAAAG AGCGAGGCTG TTTATCAGGA TGACAAGTTG GTCAGGAAAA 701CCCAATGGGA TAAGGATGGT TATTTAATCG AACCCTGA

This encodes a protein having amino acid sequence <SEQ ID 704>:

1 MKKLSRIVFS IVLLGFSAAL PAQTYSVYFN QNGKLTATMS SAAYIRQYSV 51 AAGIAHAQDFYYPSMKKYSE PYIVASTQIK SFVPTLQNGM LILWHFNGQK 101 KMAGGFSKGK PDGEWVNWYPNGKKSAVMPY KNGLSEGTGY RYYRNGGKES 151 EIQFKQNKAN GVWKQWYADG SIKTEMVMVNDEPAKILTWD ESGRLLSELS 201 IRHHKRNGVV LEWYEDGSKK SEAVYQDDKL VRKTQWDKDGYLIEP*

ORF27ng and ORF27-1 show 98.8% identity in 245 aa overlap:

Based on this analysis, including the putative leader sequence in thegonococcal protein, it was predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF27-1 (24.5 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 17A shows the results ofaffinity purification of the GST-fusion protein, and FIG. 17B shows theresults of expression of the His-fusion in E. coli. Purified GST-fusionprotein was used to immunise mice, whose sera were used for ELISA, whichgave a positive result, confirming that ORF27-1 is a surface-exposedprotein and a useful immunogen.

Example 84

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 705>:

1 ATGAAATTTA CCAAGCACCC CGTCTGGGCA ATGGCGTTCC GCCCATTTTA 51 TTCGCTGGCGGCTCTGTACG GCGCATTGTC CGTATTGCTG TGGGGTTTCG 101 GCTACACGGG AACGCACkAGCTGTCCGGTT TCTATTGGCA CGCGCATGAg 151 ATGATTTGGG GTTATGCCGG ACTGGTCGTCATCGCCTTCC TGCTGACCGC 201 CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGCGTaTCTGGTC 251 GGCTTGACTA TCTTTTGGCT GGCTGCGCGG ATTGCCGCCT TTATCCCGGG301 TTGGGGTGCG TCGGCAAGCG GCATACTCGG TACGCTGTTT TTCTGGTACG 351GCGCGGTGTG CATGGCTTTG CCCGTTATCC GTTCGCAGAA TCAACGCAAC 401 TATGTTgCCGTGTTCGCGCT GTTCGTCTTG GGCGGCACGC ATGCGGCGTT 451 CCACGTCCAG CTGCACAACGGCAACCTAGG CGGACTCTTG AGCGGATTGC 501 AGTCGGGCTT GGTGATG

This corresponds to the amino acid sequence <SEQ ID 706; ORF47>:

1 MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHX LSGFYWHAHE 51 MIWGYAGLVVIAFLLTAVAT WTGQPPTRGG VLVGLTIFWL AARIAAFIPG 101 WGASASGILG TLFFWYGAVCMALPVIRSQN QRNYVAVFAL FVLGGTHAAF 151 HVQLHNGNLG GLLSGLQSGL VM

Further work revealed the complete nucleotide sequence <SEQ ID 707>:

1 ATGAAATTTA CCAAGCACCC CGTCTGGGCA ATGGCGTTCC GCCCATTTTA 51 TTCGCTGGCGGCTCTGTACG GCGCATTGTC CGTATTGCTG TGGGGTTTCG 101 GCTACACGGG AACGCACGAGCTGTCCGGTT TCTATTGGCA CGCGCATGAG 151 ATGATTTGGG GTTATGCCGG ACTGGTCGTCATCGCCTTCC TGCTGACCGC 201 CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGCGTTCTGGTCG 251 GCTTGACTAT CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT TATCCCGGGT301 TGGGGTGCGT CGGCAAGCGG CATACTCGGT ACGCTGTTTT TCTGGTACGG 351CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TTCGCAGAAT CAACGCAACT 401 ATGTTGCCGTGTTCGCGCTG TTCGTCTTGG GCGGCACGCA TGCGGCGTTC 451 CACGTCCAGC TGCACAACGGCAACCTAGGC GGACTCTTGA GCGGATTGCA 501 GTCGGGCTTG GTGATGGTGT CGGGTTTTATCGGTCTGATT GGTACGCGGA 551 TTATTTCGTT TTTTACGTCC AAACGCTTGA ATGTGCCGCAGATTCCCAGT 601 CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTGCCCATGC TGACTGCCAT651 GCTGATGGCG CACGGTGTGT TGGCTTGGCT GTCTGCCGTT TTTGCCTTTG 701CGGCAGGTGT GATTTTTACC GTGCAGGTGT ACCGCTGGTG GTATAAACCC 751 GTGTTGAAAGAGCCGATGCT GTGGATTCTG TTTGCCGGCT ATCTGTTTAC 801 CGGATTGGGG CTGATTGCGGTCGGCGCGTC TTATTTCAAA CCCGCTTTCC 851 TCAATCTGGG TGTGCATCTG ATCGGGGTCGGCGGTATCGG CGTGCTGACT 901 TTGGGCATGA TGGCGCGTAC CGCGCTTGGT CATACGGGCAATCCGATTTA 951 TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG ATGGCGGCAA1001 CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC CTACACGCAC 1051AGCATCCGCA CCTCTTCGGT TTTGTTTGCA CTCGCGCTTT TGGTGTATGC 1101 GTGGAAGTATATTCCTTGGC TGATTCGTCC GCGTTCGGAC GGCAGGCCCG 1151 GTTGA

This corresponds to the amino acid sequence <SEQ ID 708; ORF47-1>:

1 MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE LSGFYWHAHE 51MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTIFWL AARIAAFIPG 101WGASASGILG TLFFWYGAVC MALPVIRSQN QRNYVAVFAL FVLGGTHAAF 151 HVQLHNGNLGGLLSGLQSGL VMVSGFIGLI GTRIISFFTS KRLNVPQIPS 201PKWVAQASLW LPMLTAMLMA HGVLAWLSAV FAFAAGVIFT VQVYRWWYKP 251VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL IGVGGIGVLT 301 LGMMARTALGHTGNPIYPPP KAVPVAFWLM MAATAVRMVA VFSSGTAYTH 351 SIRTSSVLFA LALLVYAWKYIPWLIRPRSD GRPG*

Computer analysis of this amino acid sequence predicts a leader peptideand also gave the following results:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF47 shows 99.4% identity over a 172aa overlap with an ORF (ORF47a)from strain A of N. meningitidis:

The complete length ORF47a nucleotide sequence <SEQ ID 709> is:

1 ATGAAATTTA CCAAGCACCC CGTTTGGGCA ATGGCGTTCC GCCCGTTTTA 51 TTCACTGGCGGCTCTGTACG GCGCATTGTC CGTATTGCTG TGGGGTTTCG 101 GCTACACGGG AACGCACGAGCTGTCCGGTT TCTATTGGCA CGCGCATGAG 151 ATGATTTGGG GTTATGCCGG ACTGGTCGTCATCGCCTTCC TGCTGACCGC 201 CGTCGCCACT TGGACGGGGC AGCCGCCCAC GCGGGGCGGCGTTCTGGTCG 251 GCTTGACTAT CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT TATCCCGGGT301 TGGGGTGCGT CGGCAAGCGG CATACTCGGT ACGCTGTTTT TCTGGTACGG 351CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TTCGCAGAAT CAACGCAATT 401 ATGTTGCCGTGTTCGCGCTG TTCGTCTTGG GCGGTACGCA CGCGGCGTTC 451 CACGTCCAGC TGCACAACGGCAACCTAGGC GGACTCTTGA GCGGATTGCA 501 GTCGGGCTTG GTGATGGTGT CGGGTTTTATCGGTCTGATT GGTACGCGGA 551 TTATTTCGTT TTTTACGTCC AAACGGTTGA ATGTGCCGCAGATTCCCAGT 601 CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTGCCCATGC TGACCGCCAT651 GCTGATGGCG CACGGCGTGA TGCCTTGGCT GTCGGCGGCT TTCGCGTTTG 701CGGCAGGTGT GATTTTTACC GTGCAGGTGT ACCGCTGGTG GTATAAGCCT 751 GTGTTGAAAGAGCCGATGCT GTGGATTCTG TTTGCCGGCT ATCTGTTTAC 801 CGGATTGGGG CTGATTGCGGTCGGCGCGTC TTATTTCAAA CCCGCTTTCC 851 TCAATCTGGG TGTGCATCTG ATCGGGGTCGGCGGTATCGG CGTGCTGACT 901 TTGGGCATGA TGGCGCGTAC CGCGCTCGGT CATACGGGCAATCCGATTTA 951 TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG ATGGCGGCAA1001 CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC CTACACGCAC 1051AGCATACGCA CCTCTTCGGT TTTGTTTGCA CTCGCGCTTT TGGTGTATGC 1101 GTGGAAGTATATTCCTTGGC TGATTCGTCC GCGTTCGGAC GGCAGGCCCG 1151 GTTGA

This encodes a protein having amino acid sequence <SEQ ID 710>:

1 MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE LSGFYWHAHE 51MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTIFWL AARIAAFIPG 101WGASASGILG TLFFWYGAVC MALPVIRSQN QRNYVAVFAL FVLGGTHAAF 151 HVQLHNGNLGGLLSGLQSGL VMVSGFIGLI GTRIISFFTS KRLNVPQIPS 201PKWVAQASLW LPMLTAMLMA HGVMPWLSAA FAFAAGVIFT VQVYRWWYKP 251VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL IGVGGIGVLT 301 LGMMARTALGHTGNPIYPPP KAVPVAFWLM MAATAVRMVA VFSSGTAYTH 351 SIRTSSVLFA LALLVYAWKYIPWLIRPRSD GRPG*

ORF47a and ORF47-1 show 99.2% identity in 384 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF47 shows 97.1% identity over 172 aa overlap with a predicted ORF(ORF47ng) from N. gonorrhoeae:

The ORF47ng nucleotide sequence <SEQ ID 711> is predicted to encode aprotein comprising amino acid sequence <SEQ ID 712>:

1 MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE LSGFYWHAHE 51MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTAFWL AARIAAFIPG 101WGAAASGILG TLFFWYGAVC MALPVIRSQN RRNYVAVFAI FVLGGTHAAF 151 HVQLHNGNLGGLLSGLQSGL VMVWGFIGLI GMKIISFFTS KRLKLPQIPS 201 PKWVAHASLW LPMLNAILMAHRVMPWLSAA FPFAAGVIFT VQVYAGGITP 251 IEETSCGSVA GICYRLGNSS G

The predicted leader peptide and transmembrane domains are identical(except for an Ile/Ala substitution at residue 87 and an Leu/Ilesubstitution at position 140) to sequences in the meningococcal protein(see also Pseudomonas stutzeri orf396, accession number e246540):

TM segments in ORF47ng INTEGRAL Likelihood = −5.63 Transmembrane 52 - 68INTEGRAL Likelihood = −3.88 Transmembrane 169 - 185 INTEGRAL Likelihood= −3.08 Transmembrane 82 - 98 INTEGRAL Likelihood = −1.91 Transmembrane134 - 150 INTEGRAL Likelihood = −1.44 Transmembrane 107 - 123 INTEGRALLikelihood = −1.38 Transmembrane 227 - 243

Further work revealed the complete gonococcal DNA sequence <SEQ ID 713>:

1 ATGAAATTTA CCAAACATCC CGTCTGGGCA ATGGCGTTCC GCCCGTTTTA 51 TTCACTGGCGGCACTGTACG GCGCATTGTC CGTATTGCTG TGGGGTTTCG 101 GCTACACGGG AACGCACGAGCTGTCCGGTT TCTATTGGCA CGCGCATGAG 151 ATGATTTGGG GTTATGCCGG TCTCGTCGTCATCGCCTTCC TGCTGACCGC 201 CGTCGCCACT TGGACGGGAC AGCCGCCCAC GAGGGGCGGCGTTCTGGTCG 251 GCTTGACCGC CTTTTGGCTG GCTGCGCGGA TTGCCGCCTT TATCCCGGGT301 TGGGGTGCGG CGGCAAGCGG CATACTCGGT ACGCTGTTTT TCTGGTACGG 351CGCGGTGTGC ATGGCTTTGC CCGTTATCCG TtcgCAAAAC CGGCGCAACT 401 ATGtcgCCGTATTCGCAATA TTTGTGCTGG GCGGTACGCA TGCGgcgTTC 451 CACGtccAgc tGCACAACGGCAACCTAGGC GGACTCTTGA GCGGATTGCA 501 GTCGGGCCTG GTTATGGTGT CGGGCTTTATCGGCCTGATT GGGATGAGGA 551 TTATTTCGTT TTTTACGTCC AAACGGTTGA ACGTGCCGCAGATTCCCAGT 601 CCGAAATGGG TGGCGCAGGC TTCGCTGTGG CTACCCATGC TGACCGCCAT651 ACTGATGGCG CACGGCGTGA TGCCTTGGCT GTCGGCGGCT TTCGCGTTTG 701CGGCGGGCGT GATTTTTACC GTACAGGTGT ACCGCTGGTG GTATAAACCC 751 GTATTGAAAGAACCGATGCT GTGGATTCTG TTTGCCGGCT ATCTGTTTAC 801 CGGATTGGGG CTGATTGCGGTCGGCGCGTC TTATTTCAAA CCTGCCTTCC 851 TCAATCTGGG CGTACATCTG ATCGGGGTCGGCGGTATCGG CGTGCTGACT 901 TTGGGCATGA TGGCGCGTAC CGCGCTCGGT CATACGGGCAATTCGATTTA 951 TCCGCCGCCC AAAGCCGTTC CCGTTGCGTT TTGGCTGATG ATGGCGGCAA1001 CCGCCGTCCG TATGGTTGCC GTATTTTCTT CCGGCACTGC CTACACGCAC 1051AGCATCCGCA CGTCTTCGGT TTTGTTTGCA CTCGCGCTGC TGGTGTATGC 1101 GTGGAAATACATTCCGTGGC TGATCCGTCC GCGTTCGGAC GGCAGGCCCG 1151 GTTGA

This encodes a protein having amino acid sequence <SEQ ID 714;ORF47ng-1>:

1 MKFTKHPVWA MAFRPFYSLA ALYGALSVLL WGFGYTGTHE LSGFYWHAHE 51MIWGYAGLVV IAFLLTAVAT WTGQPPTRGG VLVGLTAFWL AARIAAFIPG 101WGAAASGILG TLFFWYGAVC MALPVIRSQN RRNYVAVFAI FVLGGTHAAF 151 HVQLHNGNLGGLLSGLQSGL VMVSGFIGLI GMRIISFFTS KRLNVPQIPS 201PKWVAQASLW LPMLTAILMA HGVMPWLSAA FAFAAGVIFT VQVYRWWYKP 251VLKEPMLWIL FAGYLFTGLG LIAVGASYFK PAFLNLGVHL IGVGGIGVLT 301 LGMMARTALGHTGNSIYPPP KAVPVAFWLM MAATAVRMVA VFSSGTAYTH 351 SIRTSSVLFA LALLVYAWKYIPWLIRPRSD GRPG*

ORF47ng-1 and ORF47-1 show 97.4% identity in 384 aa overlap:

Furthermore, ORF47ng-1 shows significant homology to an ORF fromPseudomonas stutzeri:

gnl|PID|e246540 (Z73914) ORF396 protein [Pseudomonas stutzeri] Length =396 Score = 155 bits (389), Expect = 5e−37 Identities =121/391 (30%), Positives = 169/391 (42%), Gaps = 21/391 (5%) Query: 7PVWAMAFRPFYSLAALYGALSVLLWGFGYTGTHELSGFY-------WHAHEMIWGYAGLV 59P+W +AFRPF+   +LY  L++ LW   +TG     GF        WH HEM++G+A  + Sbjct: 14PIWRLAFRPFFLAGSLYALLAIPLWVAAWTGLWP--GFQPTGGWLAWHRHEMLFGFAMAI 71 Query:60 VIAFLLTAVATWTGQPPTRGGVLVGLTAFWLAARIAAFIPGWGAAASGILGTLFFWYGAV 119V  FLLTAV TWTGQ    G  LVGL A WLAAR+  ++ G  AA    L  LF Sbjct: 72VAGFLLTAVQTWTGQTAPSGNRLVGLAAVWLAARL-GWLFGLPAAWLAPLDLLFLVALVW 130 Query:120 CMALPVIRSQNRRNYVAVFAIFVLGGTHAAFXXXXXXXXXXXXXXXXXXXXXMVSGFIGL 179 MA  +   + +RNY  V  + ++ G                          +V+  + L Sbjct: 131MMAQMLWAVRQKRNYPIVVVLSLMLGADVLILTGLLQGNDALQRQGVLAGLWLVAALMAL 190 Query:180 IGMRIISFFTSKRLNVPQIPSP-KWVAQASLWLPMLTAILMAHGV----MPWLSAAFAFA 234IG R+I FFT + L       P  W+  A L    + A+L A GV     P L   F  A Sbjct: 191IGGRVIPFFTQRGLGKVDAVKPWVWLDVALLVGTGVIALLHAFGVAMRPQPLLGLLFV-A 249 Query:235 AGVIFTVQVYRWWYKPVLKEPMLWILFAGYLFTGLGLIAVGASYF-KPAFXXXXXXXXXX 293 GV   +++ RW+ K + K  +LW L    L+  +    +   +F   A Sbjct: 250IGVGHLLRLMRWYDKGIWKVGLLWSLHVAMLWLVVAAFGLALWHFGLLAQSSPSLHALSV 309 Query:294 XXXXXXXXXMMARTALGHTGNSIYPPPKAVPVAFWLXXXXXXXXXXXXFSSGTAYTHSIR 353         M+AR  LGHTG  +  P   +  AF L            F S       + Sbjct: 310GSMSGLILAMIARVTLGHTGRPLQLPAGIIG-AFVL---FNLGTAARVFLSVAWPVGGLW 365 Query:354 TSSVLFALALLVYAWKYIPWLIRPRSDGRPG 384  ++V + LA  +Y W+Y P L+  R DG PGSbjct: 366 LAAVCWTLAFALYVWRYAPMLVAARVDGHPG 396

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 85

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 715>:

1 ..ATGCCGTCTG AAGGTTCAGA CGGCmTCGGT GyCGGGGAAy CAGAAGyGGT 51  AGCGCATGCC CAATGAGACT TCGTGGGTTT TGAAGCGGGT GTTTTCCAAG 101  CGTCCCCAGT TGTGGTAACG GTATCCGGTG TCyAArGTCA GCTTGGGyGT 151  GATGTCGAAa CCGACACCGG CGATGACACC AAGACCyAmG CTGCTGATrC 201  TGTkGCTTTC GTGATAGGsA GGTTTGyTGG kmksAsyTTG TAyrATwkkG 251  CCTssCwsTG kAGmGCCkTk CkyTGGTkkA swGrwArTAG TCGTGGTTTy 301  TkTTyyCACC GAATGAACyT GATGTTTAAC GTGTCCGTAG GCGACGCGCG 351  CGCCGATATA GGGTTTGAAT TTATCGTTGA GTTTGAAATC GTAAATGGCG 401  GACAAGCCGA GAGAAGAAAC GGCGTGGAAG CTGCCGTTTC CCTGATGTTT 451  TGTTTGGGTT TCTTTGTAGT TGTTGTTTAT CTCTTCAGTA ACTTTTTTAG 501  TAGAAGAATT ACTTTCTTTC CATTTTCTGT AACTGGCATA ATCTGCCGCT 551  ATTCTCCAGC CGCCGAAATC ..

This corresponds to the amino acid sequence <SEQ ID 716; ORF67>:

1 ..MPSEGSDGXG XGEXEXVAHA QXDFVGFEAG VFQASPVVVT VSGVXXQLGX 51  DVETDTGDDT KTXAADXVAF VIGRFXGXXL YXXAXXXXAX XWXXXXSRGF 101  XXHRMNLMFN VSVGDARADI GFEFIVEFEI VNGGQAERRN GVEAAVSLMF 151  CLGFFVVVVY LFSNFFSRRI TFFPFSVTGI ICRYSPAAEI ..

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. gonorrhoeae

ORF67 shows 51.8% identity over 199 aa overlap with a predicted ORF(ORF67ng) from N. gonorrhoeae:

The ORF67ng nucleotide sequence <SEQ ID 717> is predicted to encode aprotein comprising amino acid sequence <SEQ ID 718>:

1 MPSETVGSIV NVGVDESVGF SPPFPSIQHF YRFHRIHRIR LFRPPGPMQL 51NRHSHGSGNL GRGVWATVLS DKFPCGQVRI PACAGMTNFE IAVLSGMTVR 101VFYCARPAPV NGGRLKMPSE GSDGIGIGES EAVAHAQRGF VGFEAGVFQA 151SPVVVAVAGV QGQAGRDVYA HARHRAEAQA AAAVAFLIGV FLRMSVRINR 201NCCVSITRVG GKSTCYFFSR IDAVSDVSVG DARTDIGFEF VVEFEIVNGG 251QAERRNGVEC AVFLMFRLLV FYVKLVAAKS FIILSFQLFY VHGIFIVVPF 301PVTGIIRGDA PAAEVVADRH PGVDGMRTDV SEIIAYRAYF VFAWSGWFRI 351 IVGNAFGGVG *

Based on the presence of a several putative transmembrane domains in thegonococcal protein, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 86

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 719>

1 ATGTTTGCTT TTTTAGAAGC CTTTTTTGTC GAATACGGTT ATGCGGCTGT 51TTTTTTTGTA TTGGTCATCT GCGGTTTCGG CGTGCCGATT CCCGAGGATT 101TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA TACCAATCCG 151CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG GGGACGGCAT 201CATGTTCGCC GCCGGACGAA TTTGGGGGCA GArArTCCTA rGGTTCArAC 251CTATTGCGsG CATCATGACG CCGrAACGTT ATGAGCAGGT TCAGGAAAAA 301TTCGACAAAT ACGGTAACTG GGTCTTATTT GTCGCCCGTT TCCTGCCCGG 351TTTGAGAACG GCCGTATTTG TTACAGCCGG TATCAGCCGC AAGGTTTCAT 401ACTTGCGTTT TATCATTATG GATGGACTGG CCGCA...

This corresponds to the amino acid sequence <SEQ ID 720; ORF78>:

1 MFAFLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG VISGMGYTNP 51HIMFAVGMLG VLVGDGIMFA AGRIWGQXXL XFXPIAXIMT PXRYEQVQEK 101FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFIIM DGLAA...

Further work revealed the complete nucleotide sequence <SEQ ID 721>:

1 ATGTTTGCTT TTTTAGAAGC CTTTTTTGTC GAATACGGTT ATGCGGCTGT 51TTTTTTTGTA TTGGTCATCT GCGGTTTCGG CGTGCCGATT CCCGAGGATT 101TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA TACCAATCCG 151CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG GGGACGGCAT 201CATGTTCGCC GCCGGACGAA TTTGGGGGCA GAAAATCCTA AGGTTCAAAC 251CTATTGCGCG CATCATGACG CCGAAACGTT ATGAGCAGGT TCAGGAAAAA 301TTCGACAAAT ACGGTAACTG GGTCTTATTT GTCGCCCGTT TCCTGCCCGG 351TTTGAGAACG GCCGTATTTG TTACAGCCGG TATCAGCCGC AAGGTTTCAT 401ACTTGCGTTT TATCATTATG GATGGACTGG CCGCACTGAT TTCCGTCCCT 451ATTTGGATTT ATCTGGGCGA ATACGGTGCG CACAACATCG ATTGGCTGAT 501GGCGAAAATG CACAGCCTGC AATCGGGTAT TTTTGTTATC TTGGGTATAG 551GTGCGACCGT TGTCGCTTGG ATTTGGTGGA AAAAACGCCA ACGTATCCAG 601TTTTACCGCA GCAAATTGAA AGAAAAGCGG GCGCAACGCA AAGCCGCCAA 651GGCAGCCAAA AAAGCCGCGC AAAGCAAACA ATAA

This corresponds to the amino acid sequence <SEQ ID 722; ORF78-1>:

1 MFAFLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG VISGMGYTNP 51HIMFAVGMLG VLVGDGIMFA AGRIWGQKIL RFKPIARIMT PKRYEQVQEK 101FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFIIM DGLAALISVP 151IWIYLGEYGA HNIDWLMAKM HSLQSGIFVI LGIGATVVAW IWWKKRQRIQ 201FYRSKLKEKR AQRKAAKAAK KAAQSKQ*

Computer analysis of this amino acid sequence predicts severaltransmembrane domains, and also gave the following results:

Homology with the dedA Homologue of H. influenzae (Accession NumberP45280)

ORF78 and the dedA homologue show 58% aa identity in 144aa overlap:

Orf78: 4 FLEAFFVEYGYAAVFFVLVICGFGVPIPEDLTLVTGGVISGM--GYTNPHIMFAVGMLGV 61FL  FF EYGY AV FVL+ICGFGVPIPED+TLV+GGVI+G+     N H+M  V M+GV DedA: 20FLIGFFTEYGYWAVLFVLIICGFGVPIPEDITLVSGGVIAGLYPENVNSHLMLLVSMIGV 79 Orf78:62 LVGDGIMFAAGRIWGQXXLXFXPIAXIMTPKRYEQVQEKFDKYGNWVLFVARFLPGLRTA 121L GD  M+  GRI+G   L F PI  I+T  R   V+EKF +YGN VLFVARFLPGLR DedA: 80LAGDSCMYWLGRIYGTKILRFRPIRRIVTLQRLRMVREKFSQYGNRVLFVARFLPGLRAP 139 Orf78:122 VFVTAGISRKVSYLRFIIMDGLAA 145 +++ +GI+R+VSY+RF+++D  AA DedA: 140IYMVSGITRRVSYVRFVLIDFCAA 163Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF78 shows 93.8% identity over a 145aa overlap with an ORF (ORF78a)from strain A of N. meningitidis:

The complete length ORF78a nucleotide sequence <SEQ ID 723> is:

1 ATGTTTGCCC TTTTGGAAGC CTTTTTTGTC GAATACGGCT ATGCGGCCGT 51GTTTTTCGTT TTGGTCATCT GCGGTTTCGG CGTGCCGATT CCCGAGGATT 101TGACCTTGGT AACAGGCGGC GTGATTTCGG GTATGGGTTA TACCAATCCG 151CATATTATGT TTGCAGTCGG TATGCTCGGC GTATTGGTCG GGGACGGCAT 201CATGTTCGCC GCCGGACGCA TCTGGGGGCA GAAAATCCTC AAGTTCAAAC 251CGATTGCGCG CATCATGACG CCGAAACGTT ACGCACAGGT TCAGGAAAAA 301TTCGACAAAT ACGGCAACTG GGTGTTATTT GTCGCTCGTT TCCTGCCCGG 351TTTGCGGACT GCCGTTTTCG TTACCGCCGG CATCAGCCGC AAAGTATCGT 401ATCTGCGCTT TCTGATTATG GACGGGCTTG CCGCGCTGAT TTCCGTGCCC 451GTTTGGATTT ACTTGGGCGA GTACGGCGCG CACAACATCG ATTGGCTGAT 501GGCGAAAATG CACAGCCTGC AATCCGGCAT CTTCATCGCA TTGGGCGTGC 551TGGCGGCGGC GCTGGCGTGG TTCTGGTGGC GCAAACGCCG ACATTATCAG 601CTTTACCGCG CACAATTGAG CGAAAAACGC GCCAAACGCA AGGCGGAAAA 651GGCAGCGAAA AAAGCGGCAC AGAAGCAGCA GTAA

This encodes a protein having amino acid sequence <SEQ ID 724>:

1 MFALLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG VISGMGYTNP 51HIMFAVGMLG VLVGDGIMFA AGRIWGQKIL KFKPIARIMT PKRYAQVQEK 101FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFLIM DGLAALISVP 151VWIYLGEYGA HNIDWLMAKM HSLQSGIFIA LGVLAAALAW FWWRKRRHYQ 201LYRAQLSEKR AKRKAEKAAK KAAQKQQ*

ORF78a and ORF78-1 show 89.0% identity in 227 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF78 shows 97.4% identity over 38 aa overlap with a predicted ORF(ORF78ng) from N. gonorrhoeae:

The ORF78ng nucleotide sequence <SEQ ID 725> is predicted to encode aprotein comprising amino acid sequence <SEQ ID 726>:

1 ..YPVLFVARFL PGLRTAVFVT AGISRKVSYL RFLIMDGLAA LISVPVWIYL 51  GEYGAHNIDW LMAKMHSLQS GIFIALGVLA AALAWFWWRK RRHYQLYRAQ 101  LSEKRAKRKA EKAAKKAAQK QQ*

Further work revealed the complete gonococcal nucleotide sequence <SEQID 727>:

1 atgtttgccc tttTggaagc CTTTTTTGTC GAAtacggCt atgcGGCCGT 51GTTTTTCGTT TTGGTCATCT GCGGTTTCGG CGTGCCGATT CCCGAAGATT 101TGACCTTGGT AACGGGCGGC GTGATTTCGG GTATGGGTTA TACCAATCCG 151CATATTATGT TTGCGGTCGG TATGCTCGGC GTGTTGGCGG GCGACGGCGT 201GATGTTTGCC GCCGGACGCA TCTGGGGGCA GAAAATCCTC AAGTTCAAAC 251CGATTGCGCG CATCATGACG CCGAAACGTT ACGCGCAGGT TCAGGAAAAA 301TTCGACAAAT ACGGCAACTG GGTTCTGTTT GTCGCCCGTT TCCTGCCGGG 351TTTGCGGACT GCCGTTTTCG TTACCGCCGG CATCAGCCGC AAAGTATCGT 401ATCTGCGCTT TCTGATTATG GACGGGCTGG CCGCGCTGAT TTCCGTGCCC 451GTTTGGATTT ACTTGGGCGA GTACGGCGCG CACAACATCG ATTGGCTGAT 501GGCGAAAATG CACAGCCTGC AATCGGGCAT CTTCATCGCA TTGGGCGTGC 551TGGCGGCGGC GCTGGCGTGG TTCTGGTGGC GCAAACGCCG ACATTATCAG 601CTTTACCGCG CACAATTGAG CGAAAAACGC GCCAAACGCA AGGCGGAAAA 651GGCAGCGAAA AAAGCGGCAC AGAAGCAGCA GTAa

This corresponds to the amino acid sequence <SEQ ID 728; ORF78ng-1>:

1 MFALLEAFFV EYGYAAVFFV LVICGFGVPI PEDLTLVTGG VISGMGYTNP 51HIMFAVGMLG VLAGDGVMFA AGRIWGQKIL KFKPIARIMT PKRYAQVQEK 101FDKYGNWVLF VARFLPGLRT AVFVTAGISR KVSYLRFLIM DGLAALISVP 151VWIYLGEYGA HNIDWLMAKM HSLQSGIFIA LGVLAAALAW FWWRKRRHYQ 201LYRAQLSEKR AKRKAEKAAK KAAQKQQ*

ORF78ng-1 and ORF78-1 show 88.1% identity in 227 aa overlap:

Furthermore, orf78ng-1 shows homology to the dedA protein from H.influenzae:

sp|P45280|YG29_HAEIN HYPOTHETICAL PROTEIN HI1629 >gi|1073983|pir||D64133dedA protein (dedA) homolog - Haemophilus influenzae (strain Rd KW20)>gi|1574476 (U32836) dedA protein (dedA) [Haemophilus influenzae]Length = 212 Score = 223 bits (563), Expect = 7e−58 Identities =108/182 (59%), Positives = 140/182 (76%), Gaps = 2/182 (1%) Query: 5LEAFFVEYGYAAVFFVLVICGFGVPIPEDLTLVTGGVISGM--GYTNPHIMFAVGMLGVL 62L  FF EYGY AV FVL+ICGFGVPIPED+TLV+GGVI+G+     N H+M  V M+GVL Sbjct: 21LIGFFTEYGYWAVLFVLIICGFGVPIPEDITLVSGGVIAGLYPENVNSHLMLLVSMIGVL 80 Query:63 AGDGVMFAAGRIWGQKILKFKPIARIMTPKRYAQVQEKFDKYGNWVLFVARFLPGLRTAV 122AGD  M+  GRI+G KIL+F+PI RI+T +R   V+EKF +YGN VLFVARFLPGLR  + Sbjct: 81AGDSCMYWLGRIYGTKILRFRPIRRIVTLQRLRMVREKFSQYGNRVLFVARFLPGLRAPI 140 Query:123 FVTAGISRKVSYLRFLIMDGLAALISVPVWIYLGEYGAHNIDWLMAKMHSLQSGIFIALG 182 +++GI+R+VSY+RF+++D  AA+ISVP+WIYLGE GA N+DWL  ++   Q  I+I +G Sbjct: 141YMVSGITRRVSYVRFVLIDFCAAIISVPIWIYLGELGAKNLDWLHTQIQKGQIVIYIFIG 200 Query:183 VL 184  L Sbjct: 201 YL 202

Based on this analysis, including the presence of putative transmembranedomains, it is predicted that these proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 87

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 729>:

1 ATGAAAAAAT TATTGGCGGC CGTGATGATG GCAGGTTTGG CAGGCGCGGT 51 TTCCGCCGCCGGAGTCCACG TTGAGGACGG CTGGGCGCGC ACCACCGTCG 101 AAGGTATGAA AATAGGCGGCGCGTTCATGA AAATCCACAA CGACGAAGCC 151 AAACAAGACT TTTTGCTCGG CGGAAGCAGCCCCGTTGCCG ACCGCGTCGA 201 AGTGCATACC CACATCAACG ACAACGGCGT GATGCGGATGCGCGAAGTCG 251 AAGGCGGCGT GCCTTTGGAA GCGAAATCCG TTACCGAACT CAAACCCGGC301 AGCTATCATG TGATGTTTAT GGGTTTGAAA AAACAATTAA AAGAGGGCGA 351TAAAATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG CAAACCGTCC 401 AACTGGAAGTCAAAATCGCG CCGATGCCGG CAATGAACCA C...

This corresponds to the amino acid sequence <SEQ ID 730; ORF79>:

1 MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKIGG AFMKIHNDEA 51 KQDFLLGGSSPVADRVEVHT HINDNGVMRM REVEGGVPLE AKSVTELKPG 101 SYHVMFMGLK KQLKEGDKIPVTLKFKNAKA QTVQLEVKIA PMPAMNH..

Further work revealed the complete nucleotide sequence <SEQ ID 731>:

1 ATGAAAAAAT TATTGGCGGC CGTGATGATG GCAGGTTTGG CAGGCGCGGT 51 TTCCGCCGCCGGAGTCCACG TTGAGGACGG CTGGGCGCGC ACCACCGTCG 101 AAGGTATGAA AATAGGCGGCGCGTTCATGA AAATCCACAA CGACGAAGCC 151 AAACAAGACT TTTTGCTCGG CGGAAGCAGCCCCGTTGCCG ACCGCGTCGA 201 AGTGCATACC CACATCAACG ACAACGGCGT GATGCGGATGCGCGAAGTCG 251 AAGGCGGCGT GCCTTTGGAA GCGAAATCCG TTACCGAACT CAAACCCGGC301 AGCTATCATG TGATGTTTAT GGGTTTGAAA AAACAATTAA AAGAGGGCGA 351TAAAATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG CAAACCGTCC 401 AACTGGAAGTCAAAATCGCG CCGATGCCGG CAATGAACCA CGGTCATCAC 451 CACGGCGAAG CGCATCAGCACTAA

This corresponds to the amino acid sequence <SEQ ID 732; ORF79-1>:

1 MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKIGG AFMKIHNDEA 51 KQDFLLGGSSPVADRVEVHT HINDNGVMRM REVEGGVPLE AKSVTELKPG 101 SYHVMFMGLK KQLKEGDKIPVTLKFKNAKA QTVQLEVKIA PMPAMNHGHH 151 HGEAHQH*

Computer analysis of this amino acid sequence revealed a putative leaderpeptide and also gave the following results:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF79 shows 94.6% identity over a 147aa overlap with an ORF (ORF79a)from strain A of N. meningitidis:

The complete length ORF79a nucleotide sequence <SEQ ID 733> is:

1 ATGAAANAAC TATTGGCAGC CGTGATGATG GCAGGTTTGG CAGGCGCGGT 51 TTCCGCCGCCGGAATCCACG TTGAGGACGG CTGGGCGCGC ACCACCGTCG 101 AAGGTATGAA AATGGGCGGCGCGTTCATGA AAATCCACAA CGACGAAGCC 151 AAACAAGACT TTTTGCTCGG CGGAAGCAGCCCTGTTGCCG ACCGCGTCGA 201 AGTGCATACC CATATCAATG ATAACGGTGT GATGCGGATGCGCGAAGTCG 251 AAGGCGGCGT GCCTTTGGAG GCGAAATCCG TTACCGAACT CAAACCCGGC301 AGCTATCATG TCATGTTTAT GGGTNTGAAA AAACAATTAA AAGANGGCGA 351CAAGATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCA CAAACCGTCC 401 AACTGGAAGTCAAAACCGCG CCGATGTCGG CAATGGACCA CGGTCATCAC 451 CACGGCGAAG CGCATCAGCACTAA

This encodes a protein having amino acid sequence <SEQ ID 734>:

1 MKXLLAAVMM AGLAGAVSAA GIHVEDGWAR TTVEGMKMGG AFMKIHNDEA 51 KQDFLLGGSSPVADRVEVHT HINDNGVMRM REVEGGVPLE AKSVTELKPG 101 SYHVMFMGXK KQLKXGDKIPVTLKFKNAKA QTVQLEVKTA PMSAMDHGHH 151 HGEAHQH*

ORF79a and ORF79-1 show 94.9% identity in 157 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF79 shows 96.1% identity over 76 aa overlap with a predicted ORF(ORF79ng) from N. gonorrhoeae:

An ORF79ng nucleotide sequence <SEQ ID 735> was predicted to encode aprotein comprising amino acid sequence <SEQ ID 736>:

1 ..INDNGVMRMR EVKGGVPLEA KSVTELKPGS YHVMFMGLKK QLKEGDKIPV 51  TLKFKNAKAQ TVQLEVKTAP MSAMNHGHHH GEAHQH*

Further work revealed the complete gonococcal DNA sequence <SEQ ID 737>:

1 ATGAAAAAAT TATTGGCAGC CGTGATGATG GCAGGTTTGG CAGGCGCGGT 51 TTccgccgCcGGagTccAtG TCGAggACGG CTGGGCGCGc accaCTGtcg 101 aaggtATgaa aatggGCGGCGCgttCATga aaATCCACAA CGACGaaGcc 151 atacaaGACt ttgtgcTCgg CGGaagcatgcccgttgccg accgcGTCGA 201 AGTGCAtaca cacATCAACG ACAACGGCGT GATGCGTATGCGCGAAGTCA 251 AAGGCGGCGT GCCTTTGGAG GCGAAATCCG TTACCGAACT CAAACCCGGC301 AGCTATCACG TGATGTTTAT GGGTTTGAAA AAACAACTGA AAGAGGGCGA 351CAAGATTCCC GTTACCCTGA AATTTAAAAA CGCCAAAGCG CAAACCGTCC 401 AACTGGAAGTCAAAACCGCG CCGATGTCGG CAATGAACCA CGGTCATCAC 451 CACGGCGAAG CGCATCAGCACTAA

This corresponds to the amino acid sequence <SEQ ID 738; ORF79ng-1>:

1 MKKLLAAVMM AGLAGAVSAA GVHVEDGWAR TTVEGMKMGG AFMKIHNDEA 51 IQDFVLGGSMPVADRVEVHT HINDNGVMRM REVKGGVPLE AKSVTELKPG 101 SYHVMFMGLK KQLKEGDKIPVTLKFKNAKA QTVQLEVKTA PMSAMNHGHH 151 HGEAHQH*

ORF79ng-1 and ORF79-1 show 95.5% identity in 157 aa overlap:

Furthermore, ORF79ng-1 shows significant homology to a protein fromAquifex aeolicus:

gi|2983695 (AE000731) putative protein [Aquifex aeolicus] Length = 151Score = 63.6 bits (152), Expect = 6e−10 Identities = 38/114 (33%),Positives = 58/114 (50%), Gaps = 1/114 (0%) Query: 24VEDGWARTTVEGMKMGGAFMKIHNDEAIQDFVLGGSMPVADRVEVHTHINDNGVMRMREV 83V+  W      G       M I N+    D+++G    +A RVE+H  + +N V +M Sbjct: 27VKHPWVMEPPPGPNTTMMGMIIVNEGDEPDYLIGAKTDIAQRVELHKTVIENDVAKMVPQ 86 Query:84 KGGVPLEAKSVTELKPGSYHVMFMGLKKQLKEGDKIPVTLKFKNAKAQTVQLEV 137+  + +  K   E K   YHVM +GLKK++KEGDK+ V L F+ +   TV+  V Sbjct: 87ER-IEIPPKGKVEFKHHGYHVMIIGLKKRIKEGDKVKVELIFEKSGKITVEAPV 139

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF79-1 (15.6 kDa) was cloned in the pET vector and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 18A shows the results ofaffinity purification of the His-fusion protein. Purified His-fusionprotein was used to immunise mice, whose sera were used for ELISA(positive result) and FACS analysis (FIG. 18B) These experiments confirmthat ORF79-1 is a surface-exposed protein, and that it is a usefulimmunogen.

Example 88

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 739>:

1 ATGACGGTAA CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG CGTTAAAAAA 51 ATATCTGATTACGGGCATTT TGGTCTGGCT GCCGATTGCG GTAACGGTTT 101 GGGTGGTTTC CTATATCGTTTCCGCGTCCG ATCAGCTCGT CAACCTGCTG 151 CCGAAGCAAT GGCGGCCGCA ATATGTTTTGGGGTTTAATA TCCCGGGGCT 201 GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGATTGTTTGCCG 251 CCAACGTATT GGGTCGGCAG ATCCTCGCCG CGTGGGACAG CCTGTTGGGG301 CGGATTCCGG TTGTGAAAtC CATCTATTCG AGTGTGAAAA AAGTATCCGA 351ATacgTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG GTACTCGTGC 401 CGTTTCCCCAGCCCGGTATT TGGACGATyG CTTTCGTGTC AGGGCAGGTG 451 TCGAATGCGG TTAAGGCCGCATTGCCGAAs GACGGCGATT ATCTTTCCGT 501 GTATGTTCCG ACCACGCCGA ATCCGACCGGCGGTTACTAT ATTATGGTAA 551 AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGAAsCATTGAAA 601 TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC CCGTCAAAAC651 ATTGGCAsGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC GAACAACAAT 701 AA

This corresponds to the amino acid sequence <SEQ ID 740; ORF98>:

1 MTVTAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV SASDQLVNLL 51 PKQWRPQYVLGFNIPGLGVI VAIAVLFVTG LFAANVLGRQ ILAAWDSLLG 101 RIPVVKSIYS SVKKVSEYVLSDSSRSFKTP VLVPFPQPGI WTIAFVSGQV 151 SNAVKAALPX DGDYLSVYVP TTPNPTGGYYIMVKKSDVRE LDMSVDEXLK 201 YVISLGMVIP DDLPVKTLAX PMPSEKADLP EQQ*

Further work revealed the complete nucleotide sequence <SEQ ID 741>:

1 ATGACGGAAC nTGCGGCCGA AGGCGGCAAA GCTGCCAArG CGTTAAAAAA 51 ATATCTGATTACGGGCATTT TGGTCTGGCT GCCGATTGCG GTAACGGTTT 101 GGGTGGTTTC CTATATCGTTTCCGCGTCCG ATCAGCTCGT CAACCTGCTG 151 CCGAAGCAAT GGCGGCCGCA ATATGTTTTGGGGTTTAATA TCCCGGGGCT 201 GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGATTGTTTGCCG 251 CCAACGTATT GGGTCGGCAG ATCCTCGCCG CGTGGGACAG CCTGTTGGGG301 CGGATTCCGG TTGTGAAATC CATCTATTCG AGTGTGAAAA AAGTATCCGA 351ATCGCTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG GTACTCGTGC 401 CGTTTCCCCAGCCCGGTATT TGGACGATTG CTTTCGTGTC AGGGCAGGTG 451 TCGAATGCGG TTAAGGCCGCATTGCCGAAG GACGGCGATT ATCTTTCCGT 501 GTATGTTCCG ACCACGCCGA ATCCGACCGGCGGTTACTAT ATTATGGTAA 551 AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGAAGCATTGAAA 601 TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC CCGTCAAAAC651 ATTGGCAGGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC GAACAACAAT 701 AA

This corresponds to the amino acid sequence <SEQ ID 742; ORF98-1>:

1 MTEXAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV SASDQLVNLL 51 PKQWRPQYVLGFNIPGLGVI VAIAVLFVTG LFAANVLGRQ ILAAWDSLLG 101 RIPVVKSIYS SVKKVSESLLSDSSRSFKTP VLVPFPQPGI WTIAFVSGQV 151 SNAVKAALPK DGDYLSVYVP TTPNPTGGYYIMVKKSDVRE LDMSVDEALK 201 YVISLGMVIP DDLPVKTLAG PMPSEKADLP EQQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF98 shows 96.1% identity over a 233aa overlap with an ORF (ORF98a)from strain A of N. meningitidis:

The complete length ORF98a nucleotide sequence <SEQ ID 743> is:

1 ATGACGGAAC CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG CGTTAAAAAA 51 ATATCTGATTACGGGCATTT TGGTCTGGCT GCCGATTGCG GTAACGGTTT 101 GGGTGGTTTC CTATATCGTTTCCGCGTCCG ATCAGCTCGT CAACCTGCTG 151 CCGAAGCAAT GGCGGCCGCA ATATGTTTTGGGGTTTAATA TCCCGGGGCT 201 GGGCGTTATC GTTGCCATTG CCGTATTGTT TGTAACCGGATTATTTGCCG 251 CAAACGTATT GGGCCGGCAG ATTCTTGCCG CGTGGGACAG CTTGTTGGGG301 CGGATTCCGG TTGTGAAGTC CATCTATTCG AGTGTGAAAA AAGTATCCGA 351NTCGTTGCTG TCCGACAGCA GCCGTTCGTT TAAAACACCA GTACTCGTGC 401 CGTTTCCCCAATCGGGTATT TGGACAATCG CATTCGTGTC CGGTCAGGTG 451 TCGAATGCGG TTAAGGCCGCATTGCCGAAG GACGGCGATT ATCTTTCCGT 501 GTATGTTCCG ACCACGCCGA ATCCGACCGGCGGTTACTAT ATTATGGTAA 551 AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGAAGCGTTGAAA 601 TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC CCGTCAAAAC651 ATTGGCAGGA CCTATGCCGT CTGAAAAGGC GGATTTGCCC GAACAACAAT 701 AA

This encodes a protein having amino acid sequence <SEQ ID 744>:

1 MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV SASDQLVNLL 51PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFAANVLGRQ ILAAWDSLLG 101RIPVVKSIYS SVKKVSXSLL SDSSRSFKTP VLVPFPQSGI WTIAFVSGQV 151SNAVKAALPK DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE LDMSVDEALK 201YVISLGMVIP DDLPVKTLAG PMPSEKADLP EQQ*

ORF98a and ORF98-1 show 98.7% identity in 233 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF98 shows 95.3% identity over a 233 aa overlap with a predicted ORF(ORF98ng) from N. gonorrhoeae:

The complete length ORF98ng nucleotide sequence <SEQ ID 745> ispredicted to encode a protein having amino acid sequence <SEQ ID 746>:

1 MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV SASDQLVNLL 51PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFAANVLGRQ ILAAWDSLLX 101RIPVVKSIYS SVKKVSESLL SDSSRSFKTP VLVPFPQSGI WTIAFVSGQV 151SNAVKAALPQ DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE LDMSVDEALK 201YVISLGMVIP DDLPVKTLAG PMPPEKAELP EQQ*

Further work revealed the complete nucleotide sequence <SEQ ID 747>:

1 ATGACGGAAC CTGCGGCCGA AGGCGGCAAA GCTGCCAAGG CGTTAAAAAA 51ATATCTGATT ACAGGCATTT TGGTCTGGCT GCCGATTGCG GTAACGGTTT 101GGGTGGTTTC CTATATCGTT TCCGCGTCCG ACCAGCTTGT CAACCTGCTG 151CCGAAGCAAT GGCGGCCGCA ATATGTTTTG GGGTTTAATA TCCCCGGGCT 201CGGCGTTATT GTTGCCATTG CCGTATTGTT TGTAACCGGA TTATTTGCCG 251CAAACGTGTT GGGCCGGCAG ATTCTTGCCG CGTGGGACAG CCTGTTgggg 301cggaTTCCGG TTGTCAAATC CATCTATTCG AGTGTGAAAA AAGTATCCGA 351ATCGCTGCTG TCCGACAGCA GCCGTTCGTT TAAAACGCCG GTACTCGTGC 401CGTTTCCCCA ATCGGGTATT TGGACAATCG CATTCGTGTC CGGTCAGGTG 451TCGAATGCGG TTAAGGCCGC ATTGCCGCAG GATGGCGATT ATCTTTCCGT 501GTATGTCCCG ACCACGCCCA ACCCGACCGG CGGTTACTAT ATTATGGTAA 551AGAAAAGCGA TGTGCGCGAA CTCGATATGA GCGTGGACGA AGCGTTGAAA 601TATGTGATTT CGCTGGGTAT GGTCATCCCT GACGACCTGC CCGTCAAAAC 651ATTGGCAGGA CCTATGCCGC CTGAAAAGGC GGAGTTGCCC GAACAACAAT 701 AA

This corresponds to the amino acid sequence <SEQ ID 748; ORF98ng-1>:

1 MTEPAAEGGK AAKALKKYLI TGILVWLPIA VTVWVVSYIV SASDQLVNLL 51PKQWRPQYVL GFNIPGLGVI VAIAVLFVTG LFAANVLGRQ ILAAWDSLLG 101RIPVVKSIYS SVKKVSESLL SDSSRSFKTP VLVPFPQSGI WTIAFVSGQV 151SNAVKAALPQ DGDYLSVYVP TTPNPTGGYY IMVKKSDVRE LDMSVDEALK 201YVISLGMVIP DDLPVKTLAG PMPPEKAELP EQQ*

ORF98ng-1 and ORF98-1 show 97.9% identity in 233 aa overlap:

Based on this analysis, including the fact that the putativetransmembrane domains in the gonococcal protein are identical to thesequences in the meningococcal protein, it is predicted that theproteins from N. meningitidis and N. gonorrhoeae, and their epitopes,could be useful antigens for vaccines or diagnostics, or for raisingantibodies.

Example 89

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 749>:

1 ATgAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG CCGTCGGACT 51GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC GTACTCGGAC 101AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG TTCGCTGATT 151GCCGTCGTGG TGTGGTATTT CTTGTTTAAA TTCATTATCG GsGgTACTCA 201ATATCCCCGA AAAGATGCAG CGTTTCGGTT CGGCnCGTAA AGGCCkCAAG 251ssCGsGCTTG CCTTGAACAA GGCGGGTTTG GCGTATTTTG AAGGGCGTTT 301TGAAAAGGCG GAACTAGAAG CCTCACGCGT GTTGGTCAAC AAAGtAGGCC 351GaGAGACAAC CGGACTTTGG CATTGATGCT GrGCGCGCAC GCCGCCGGAC 401AGATGGAAAA CATCGAssTG CGCGACCGTT ATCTTGCGGA AATCGCCAAA 451CTGCCGGAAA AACAGCAGCT TTCCCGTTAT CTTTTGTTGG CGGAATCGGC 501GTTGAACCGG CGCGATTACG AAGCGGCGGA AGCCAATCTT CATGCGGCGG 551CGAAGATGAA TGCCAACCTT ACGCGCCTCG TGCGTCTGCA .ATTCGTTAC 601GCTTTCGACA GGGGCGACGC GTTGCAGGTT CTGGCAAAAA CCGAAAAACT 651TTCCAAGGCG GGCGCGTTGG GCAAATCGGA AATGGAACGG TATCAAAATT 701GGGCATATCC GTCGCCAGCT GGCGGATGCT GCCGATGCCG CCGCTTTGAA 751AACCTGCCTG AAGCGGATTC CCGACAGCCT CAAAAACGGG GAATTGAGCG 801TATCGGTTGC GGAAAAGTAC GAACGTTTGG GACTGTATGC CGATGCGGTC 851AAATGGGTCA AACAGCATTA TCCGCAsAAC CGCCGCCCCG AGCTTTTGGA 901AGCCTTTGTC GAAAGCGTGC GCTTTTTGGG CGAGCGCGAA CAGCAGAAAG 951CCATCGATTT TGCCGATGCT TGGCTGAAAG AACAGCCCGA TAACGCGCTT 1001CTGCTGATGT ATCTCGGTCG GCTCGCCTTC GGCCGCAAAC TTTGGGGCAA 1051GGCAAAAGGC TACCTTGAAG CGAGCATTGC ATTAAAGCCG AGTATTTCCG 1101CGCGTTTGGT TCTAACAAAG GTTTTCGACG AAATCGGAGA ACCGCAGAAG 1151GCGGAGGCGC AC...

This corresponds to the amino acid sequence <SEQ ID 750; ORF100>:

1 MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN LHAFVLGSLI 51AVVVWYFLFK FIIGVLNIPE KMQRFGSARK GXKXXLALNK AGLAYFEGRF 101EKAELEASRV LVNKVGRDNR TLALMLXAHA AGQMENIXXR DRYLAEIAKL 151PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT RLVRLXIRYA 201FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQLA DAADAAALKT 251CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP XNRRPELLEA 301FVESVRFLGE REQQKAIDFA DAWLKEQPDN ALLLMYLGRL AFGRKLWGKA 351KGYLEASIAL KPSISARLVL TKVFDEIGEP QKAEAH...

Further work revealed the complete nucleotide sequence <SEQ ID 751>:

1 ATGAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG CCGTCGGACT 51GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC GTACTCGGAC 101AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG TTCGCTGATT 151GCCGTCGTGG TGTGGTATTT CTTGTTTAAA TTCATTATCG GCGTACTCAA 201TATCCCCGAA AAGATGCAGC GTTTCGGTTC GGCGCGTAAA GGCCGCAAGG 251CCGCGCTTGC CTTGAACAAG GCGGGTTTGG CGTATTTTGA AGGGCGTTTT 301GAAAAGGCGG AACTAGAAGC CTCACGCGTG TTGGTCAACA AAGAGGCCGG 351AGACAACCGG ACTTTGGCAT TGATGCTGGG CGCGCACGCC GCCGGACAGA 401TGGAAAACAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT CGCCAAACTG 451CCGGAAAAAC AGCAGCTTTC CCGTTATCTT TTGTTGGCGG AATCGGCGTT 501GAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT GCGGCGGCGA 551AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT TCGTTACGCT 601TTCGACAGGG GCGACGCGTT GCAGGTTCTG GCAAAAACCG AAAAACTTTC 651CAAGGCGGGC GCGTTGGGCA AATCGGAAAT GGAACGGTAT CAAAATTGGG 701CATACCGCCG CCAGCTGGCG GATGCTGCCG ATGCCGCCGC TTTGAAAACC 751TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT TGAGCGTATC 801GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT GCGGTCAAAT 851GGGTCAAACA GCATTATCCG CACAACCGCC GCCCCGAGCT TTTGGAAGCC 901TTTGTCGAAA GCGTGCGCTT TTTGGGCGAG CGCGAACAGC AGAAAGCCAT 951CGATTTTGCC GATGCTTGGC TGAAAGAACA GCCCGATAAC GCGCTTCTGC 1001TGATGTATCT CGGTCGGCTC GCCTACGGCC GCAAACTTTG GGGCAAGGCA 1051AAAGGCTACC TTGAAGCGAG CATTGCATTA AAGCCGAGTA TTTCCGCGCG 1101TTTGGTTCTA GCAAAGGTTT TCGACGAAAT CGGAGAACCG CAGAAGGCGG 1151AGGCGCAGCG CAACTTGGTT TTGGAAGCCG TCTCCGATGA CGAACGTCAC 1201GCAGCGTTAG AGCAGCATAG CTGA

This corresponds to the amino acid sequence <SEQ ID 752; ORF100-1>:

1 MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN LHAFVLGSLI 51AVVVWYFLFK FIIGVLNIPE KMQRFGSARK GRKAALALNK AGLAYFEGRF 101EKAELEASRV LVNKEAGDNR TLALMLGAHA AGQMENIELR DRYLAEIAKL 151PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT RLVRLQLRYA 201FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQLA DAADAAALKT 251CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP HNRRPELLEA 301FVESVRFLGE REQQKAIDFA DAWLKEQPDN ALLLMYLGRL AYGRKLWGKA 351KGYLEASIAL KPSISARLVL AKVFDEIGEP QKAEAQRNLV LEAVSDDERH 401 AALEQHS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF100 shows 93.5% identity over a 386aa overlap with an ORF (ORF100a)from strain A of N. meningitidis:

The complete length ORF100a nucleotide sequence <SEQ ID 753> is:

1 ATGAAAACGG TAGTCTGGAT TGTCGTCCTG TTTGCCGCCG CNNTCGGGCT 51GGCATTGGCG TCGGGCATTN ACACCGGCGA CGTGTATATC GTACTCGGAC 101AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG TTCGCTGATT 151GCCGTCGTGG TGTGGTATTT CCTGTTCAAA TTCATCATCG GCGTACTCAA 201TANCCCCGAA AAGATGCAGC GTTTCGGTTC GGCGCGTAAA GGCCGCAAGG 251CCGCGCTTGC TTTGAACAAG GCGGGTTTGG CGTATTTTGA AGGGCGTTTT 301GAAAAGGCGG AACTTGAAGC CTCGCGCGTA TTGGGAAACA AAGAGGCGGG 351GGATAACCGG ACTTTGGCAT TGATGTTGGG CGCACATGCC GCCGGGCAGA 401TGGAAAACAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT CGCCAAACTG 451CCGGAAAAGC AGCAGCTTTC CCGTTATCTT TTGTTGGCGG AATCGGCGTT 501GAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT GCGGCGGCGA 551AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT TCGTTACGCT 601TTCGACAGGG GCGACGCGTT GCAGGTTCTG GCAAAAACCG AAAAANTTTC 651CAAGGCGGGC GCGTNGGGCA AATCGGAAAT GGAACGGTAT CAAAATTGGG 701CATACCGCCG CCAGCTGNCG GATGCTGCCG ATGCCGCCGC TTTGAAAACC 751TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT TGAGCGTATC 801GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT GCGGTCAAAT 851GGGTCAAACA GCATTATCCG CACAACCGCC GACCCGAACT TTTGGAAGCN 901TTTGTCGAAA GCGTGCGCTT TTTGGGCGAA CGCGATCAGC AGAAAGCCAT 951CGATTTTGCC GATGCTTGGC TGAAAGAACA GCCCGATAAT GCGCTTCTGC 1001TGANGTATCT CGGTCGGCTC GCCTACGGCC GCAAACTTTG GGGCAAGGCA 1051AAAGGCTACC TTGAAGCGAG CATTGCATTA AAGCCGAGTA TTTCCGCGCG 1101TTTGGTTCTG GCAAAGGTTT TTGACGAAAC CGGAGAACCG CAGAAGGCGG 1151AGGCGCAGCG CAACTTGGTT TTGGCAAGCG TTGCCGAGGA AAACCGNCCT 1201TCCGCCGAAA CCCATTGA

This encodes a protein having amino acid sequence <SEQ ID 754>:

1 MKTVVWIVVL FAAAXGLALA SGIXTGDVYI VLGQTMLRIN LHAFVLGSLI 51AVVVWYFLFK FIIGVLNXPE KMQRFGSARK GRKAALALNK AGLAYFEGRF 101EKAELEASRV LGNKEAGDNR TLALMLGAHA AGQMENIELR DRYLAEIAKL 151PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT RLVRLQLRYA 201FDRGDALQVL AKTEKXSKAG AXGKSEMERY QNWAYRRQLX DAADAAALKT 251CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP HNRRPELLEA 301FVESVRFLGE RDQQKAIDFA DAWLKEQPDN ALLLXYLGRL AYGRKLWGKA 351KGYLEASIAL KPSISARLVL AKVFDETGEP QKAEAQRNLV LASVAEENRP 401 SAETH*

ORF100a and ORF100-1 show 95.1% identity in 406 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF100 shows 93.3% identity over a 386 aa overlap with a predicted ORF(ORF100ng) from N. gonorrhoeae:

The complete length ORF100ng nucleotide sequence <SEQ ID 755> is:

1 ATGAAAACGG TAGTCTGGAT TGTTGTCCTG TTTGCCGCCG CCGTCGGACT 51GGCGCTGGCT TCGGGCATTT ACACCGGCGA CGTGTATATC GTACTCGGAC 101AGACCATGCT CAGAATCAAC CTGCACGCCT TTGTGTTAGG TTCGCTGATT 151GCCGTCGTGG TGTGGTATTT CCTGTTTAAA TTCATCATCG GCGTACTCAA 201TATCCCCGAA AATATGCGGC GTTCCGGTTC GGCGCGGAAA GGCCGCAAGG 251CCGCGCTTGC CTTGAATAAG GCGGGTTTGG CGTATTTCGA AGGGCGTTTT 301GAAAAGGCGG AACTCGAAGC CTCTCGAGTG TTGGGCAACA AAGAGGCCGG 351AGACAACCGG ACTTTGGCAT TGATGCTGGG CGCGCACGCG GCAGGACAGA 401TGGAAAATAT CGAGCTGCGC GACCGTTATC TTGCGGAAAT CGCCAAACTG 451CCGGAAAAAC AGCAGCTTTC CCGCTATCTT CTGCTGGCGG AATCGGCGTT 501AAACCGGCGC GATTACGAAG CGGCGGAAGC CAATCTTCAT GCGGCGGCGA 551AGATGAATGC CAACCTTACG CGCCTCGTGC GTCTGCAACT TCGTTACGCC 601TTCGATCGGG GCGATGCGTT GCAGGTTCTG GCAAAAaccG AAAAACTTTC 651CAAGGCGGGC GCGTTGGGCA AATCGGAAAT GGAACGGTAT CAAAATTGGG 701CATACCGCCG CCAGATGGCG GATGCTGCCG ATGCCGCCGC TTTGAAAACC 751TGCCTGAAGC GGATTCCCGA CAGCCTCAAA AACGGGGAAT TGagcGTATC 801GGTTGCGGAA AAGTACGAAC GTTTGGGACT GTATGCCGAT GCGGTCAAAT 851GGGTCAAACA GCATTATCCG CACAACCGCC GCCCCGAGCT TTTGGAAGCC 901TTTGTCGAAA GCGTGCGCTT TTTGGGCGAG CGCGAACAGC AGAAAGCCAT 951CGATTTTGCC GATTCTTGGC TGAAAGAACA GCCCGATAAC GCGCTTCTGC 1001TGATGTATCT CGGCCGGCTC GCCTACGGCC GCAAACTTTG GGGTAAGGCA 1051AAAGGCTACC TTGAAGCGAG TATTGCACTG AAGCCGAGTA TTCCGGCGCG 1101TTTGGTGTTG GCAAAGGTTT TTGACGAAAC CGCACAGTCG CAAAAAGCCG 1151AAGCACAGCG CAACTTGGTT TTGGCAAGCG TTGCCGGGGA AAACCGCCCT 1201TCCGCCGAAA CCCGTTGA

This encodes a protein having amino acid sequence <SEQ ID 756>:

1 MKTVVWIVVL FAAAVGLALA SGIYTGDVYI VLGQTMLRIN LHAFVLGSLI 51AVVVWYFLFK FIIGVLNIPE NMRRSGSARK GRKAALALNK AGLAYFEGRF 101EKAELEASRV LGNKEAGDNR TLALMLGAHA AGQMENIELR DRYLAEIAKL 151PEKQQLSRYL LLAESALNRR DYEAAEANLH AAAKMNANLT RLVRLQLRYA 201FDRGDALQVL AKTEKLSKAG ALGKSEMERY QNWAYRRQMA DAADAAALKT 251CLKRIPDSLK NGELSVSVAE KYERLGLYAD AVKWVKQHYP HNRRPELLEA 301FVESVRFLGE REQQKAIDFA DSWLKEQPDN ALLLMYLGRL AYGRKLWGKA 351KGYLEASIAL KPSIPARLVL AKVFDETAQS QKAEAQRNLV LASVAGENRP 401 SAETR*

ORF100ng and ORF100-1 show 95.3% identity in 402 aa overlap:

Based on this analysis, including the presence of a putative leadersequence, a putative transmembrane domain, and a RGD motif, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 90

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 757>

1 ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG TCATTTCGTG 51GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT ATGGCGATGA 101TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC GGGCATGGCG 151GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG CGGTCGTGTT 201CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC GGCTGGGTAC 251ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA GTTGTATTGC 301GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT TTTCACACCG 351CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG GTTGCCGCGC 401TGTATsTGGT CGTGTTCAAA CCGTTTTGA

This corresponds to the amino acid sequence <SEQ ID 758; ORF102>:

1 MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN PEYVRLSGMA 51VRLYRFMSPL GFGAVVFGAA IPFAAGWWGS GWVHVKLCLG LMLLAYQLYC 101GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYXVVFK PF*

Further work revealed the complete nucleotide sequence <SEQ ID 759>:

1 ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG TCATTTCGTG 51GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT ATGGCGATGA 101TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC GGGCATGGCG 151GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG CGGTCGTGTT 201CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC GGCTGGGTAC 251ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA GTTGTATTGC 301GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT TTTCACACCG 351CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG GTTGCCGCGC 401TGTATCTGGT CGTGTTCAAA CCGTTTTGA

This corresponds to the amino acid sequence <SEQ ID 760; ORF102-1>:

1 MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN PEYVRLSGMA 51VRLYRFMSPL GFGAVVFGAA IPFAAGWWGS GWVHVKLCLG LMLLAYQLYC 101GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK PF*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with HP1484 Hypothetical Integral Membrane Protein of H. pylori(Accession Number AE000647)

ORF102 and HP1484 show 33% aa identity in 143aa overlap:

orf102 3 FSWFKLFHLFFVISWFAGLFYLPRIFVNMAMIDVPRGNPEYVRLSGMAVRLYRFMSPLGF 62F W K FH+  VISW A LFYLPR+FV  A     +     V++     +LY F++ HP1484 8FLWVKAFHVIAVISWMAALFYLPRLFVYHAENAHKKEFVGVVQIQEK--KLYSFIASPAM 65 orf10263 GAVVFGAAIPFAAG---WWGSGWVHVKLCLGLMLLAYQLYCGVLLRRFQDYSNAFSHRWY 119 G  +   +        +   GW+H KL L ++LLAY  YC   +R  +      + R+Y HP1484 66GFTLITGILMLLIEPTLFKSGGWLHAKLALVVLLLAYHFYCKKCMRELEKDPTRRNARFY 125 orf102120 RVFNEIPXXXXXXXXXXXXFKPF 142 RVFNE P             KPF HP1484 126RVFNEAPTILMILIVILVVVKPF 148Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF102 shows 99.3% identity over a 142aa overlap with an ORF (ORF102a)from strain A of N. meningitidis:

The complete length ORF102a nucleotide sequence <SEQ ID 761> is:

1 ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG TCATTTCGTG 51GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT ATGGCGATGA 101TTGATGTGCC GCGCGGCAAT CCCGAGTATG TGCGTCTGTC GGGCATGGCG 151GTGCGGCTGT ACCGTTTTAT GTCGCCGTTG GGCTTCGGCG CGGTCGTGTT 201CGGCGCGGCG ATACCGTTTG CCGCCGGCTG GTGGGGCAGC GGCTGGGTAC 251ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTACCA GTTGTATTGC 301GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT TTTCACACCG 351CTGGTACCGC GTGTTCAACG AAATCCCCGT GCTGCTGATG GTTGCCGCGC 401TGTATCTGGT CGTGTTCAAA CCGTTTTGA

This encodes a protein having amino acid sequence <SEQ ID 762>:

1 MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDVPRGN PEYVRLSGMA 51VRLYRFMSPL GFGAVVFGAA IPFAAGWWGS GWVHVKLCLG LMLLAYQLYC 101GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK PF*

ORF102a and ORF102-1 show complete identity in 142 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF102 shows 97.9% identity over a 142 aa overlap with a predicted ORF(ORF102ng) from N. gonorrhoeae:

The complete length ORF102ng nucleotide sequence <SEQ ID 763> is:

1 ATGATGTTTT CTTGGTTCAA GCTGTTTCAC TTGTTTTTTG TCATTTCGTG 51GTTTGCAGGG CTGTTTTACC TGCCGAGGAT TTTCGTCAAT ATGGCGATGA 101TTGATGCGCC GCGCGGCAAT CCCGAGTATG TGCGCCTGTC GGGGATGGCG 151GTGCGGTTGT ACCGTTTTAT GTCGCCTTTG GGTTTCGGCG CGGTCGTGTT 201CGGCGCGGCG ATACCGTTTG CCGCcggccg GTGGGGCagc ggctggGTTC 251ACGTCAAACT GTGTTTGGGC TTGATGCTCT TGGCTTATCA GTTGTATTGC 301GGCGTGCTGC TGCGCCGTTT TCAGGATTAC AGCAATGCTT TTTCACACCG 351CTGGTACCGC GTGTTCAAcg aAATCCCCGT GCTGCTGATG GTTGCCGCGC 401TGTATCTGGT CGTGTTCAAA CCGTTTTGA

This encodes a protein having amino acid sequence <SEQ ID 764>:

1 MMFSWFKLFH LFFVISWFAG LFYLPRIFVN MAMIDAPRGN PEYVRLSGMA 51VRLYRFMSPL GFGAVVFGAA IPFAAGRWGS GWVHVKLCLG LMLLAYQLYC 101GVLLRRFQDY SNAFSHRWYR VFNEIPVLLM VAALYLVVFK PF*

ORF102ng and ORF102-1 show 98.6% identity in 142 aa overlap:

In addition, ORF102ng shows significant homology to a membrane proteinfrom H. pylori:

gi|2314656 (AE000647) conserved hypothetical integral membrane protein[Helicobacter pylori] Length = 148 Score = 79.2 bits (192), Expect =1e−14 Identities = 50/147 (34%), Positives = 68/147 (46%), Gaps =13/147 (8%) Query: 3FSWFKLFHLFFVISWFAGLFYLPRIFVNMAMIDAPRGNPEYVRLSGMAVRLYRFMSPLGF 62F W K FH+  VISW A LFYLPR+FV  A     +     V++     +LY F++ Sbjct: 8FLWVKAFHVIAVISWMAALFYLPRLFVYHAENAHKKEFVGVVQIQEK--KLYSFIASPAM 65 Query:63 GAVVFGAAIP-------FAAGRWGSGWVHVKLCLGLMLLAYQLYCGVLLRRFQDYSNAFS 115 G  +   +        F +G    GW+H KL L ++LLAY  YC   +R  +      + Sbjct: 66GFTLITGILMLLIEPTLFKSG----GWLHAKLALVVLLLAYHFYCKKCMRELEKDPTRRN 121 Query:116 HRWYRVFNEIPXXXXXXXXXXXXFKPF 142  R+YRVFNE P             KPF Sbjct:122 ARFYRVFNEAPTILMILIVILVVVKPF 148

Based on this analysis, it is predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 91

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 765>:

  1  ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG CGGCGGCAGC 51  GGTTTGGGGC GGATGGTCTT AACTGAAGCC CGAGCCGCAC GTGCTTGATA101  TTACGGAAAC GGTCAGGCGC GGC // .....//.. ATTTCGTTTA CGATTTTGTC CGAACCGGAT ACGCCGATTA AGGCGAAGCT 51  CGACAGCGTC GACCCCGGGC TGACCACGAT GTCGTCGGGC GGTTACAACA101  GCAGTACGGA TACGGCTTCC AATGCGGTCT ACTATTATGC CCGTTCGTTT151  GTGCCGAATC CGGACGGCAA ACTCGCCACG GGGATGACGA CGCAGAATAC201  GGTTGAAATC GACGGCGTGA AAAATGTGCT GATTATTCCG TCGCTGACCG251  TGAAAAATCG CGGCGGCAAG GCGTTTGTGC GCGTGTTGGG TGCGGACGGC301  AAGGCGGCGG AACGCGAAAT CCGGACCGGT ATGAGAGACA GTATGAATAC351  CGAAGTAAAA AGCGGGTTGA AAGAGGGGGA CAAAGTGGTC ATCTCCGAAA401  TAACCGCCGC CGAGCAACAG GAAAGCGGCG AACGCGCCCT AGGCGGCCCG451  CCGCGCCGAT AA

This corresponds to the amino acid sequence <SEQ ID 766; ORF85>:

1 MAKMMKWAAV AAVAAAAVWG GWS.LKPEPH VLDITETVRR G......... 51.......... .......... .......... .......... .......... 101.......... .......... .......... .......... .......... 151.......... .......... .......... .......... .......... 201.......... .......... .......... .........I SFTILSEPDT 251PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV PNPDGKLATG 301MTTQNTVEID GVKNVLIIPS LTVKNRGGKA FVRVLGADGK AAEREIRTGM 351RDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP RR*

Further work revealed the further partial nucleotide sequence <SEQ ID767>:

1 ..GTATCGGTCG GCGCGCAGGC ATCGGGGCAG ATTAAGATAC TTTATGTCAA 51  ACTCGGGCAA CAGGTTAAAA AGGGCGATTT GATTGCGGAA ATCAATTCGA 101  CCTCGCAGAC CAATACGCTC AATACGGAAA AATCCAAGTT GGAAACGTAT 151  CAGGCGAAGC TGGTGTCGGC ACAGATTGCA TTGGGCAGCG CGGAGAAGAA 201  ATATAAGCGT CAGGCGGCGT TATGGAAGGA AAACGCGACT TCCAAAGAGG 251  ATTTGGAAAG CGCGCAGGAT GCGTTTGCCG CCGCCAAAGC CAATGTTGCC 301  GAGCTGAAGG CTTTAATCAG ACAGAGCAAA ATTTCCATCA ATACCGCCGA 351  GTCGGAATTG GGCTACACGC GCATTACCGC AACGATGGAC GGCACGGTGG 401  TGGCGATTCT CGTGGAAGAG GGGCAGACTG TGAACGCGGC GCAGTCTACG 451  CCGACGATTG TCCAATTGGC GAATCTGGAT ATGATGTTGA ACAAAATGCA 501  GATTGCCGAG GGCGATATTA CCAAGGTGAA GGCGGGGCAG GATATTTCGT 551  TTACGATTTT GTCCGAACCG GATACGCCGA TTAAGGCGAA GCTCGACAGC 601  GTCGACCCCG GGCTGACCAC GATGTCGTCG GGCGGTTACA ACAGCAGTAC 651  GGATACGGCT TCCAATGCGG TCTACTATTA TGCCCGTTCG TTTGTGCCGA 701  ATCCGGACGG CAAACTCGCC ACGGGGATGA CGACGCAGAA TACGGTTGAA 751  ATCGACGGCG TGAAAAATGT GCTGATTATT CCGTCGCTGA CCGTGAAAAA 801  TCGCGGCGGC AAGGCGTTTG TGCGCGTGTT GGGTGCGGAC GGCAAGGCGG 851  CGGAACGCGA AATCCGGACC GGTATGAGAG ACAGTATGAA TACCGAAGTA 901  AAAAGCGGGT TGAAAGAGGG GGACAAAGTG GTCATCTCCG AAATAACCGC 951  CGCCGAGCAA CAGGAAAGCG GCGAACGCGC CCTAGGCGGC CCGCCGCGCC 1001   GATAA

This corresponds to the amino acid sequence <SEQ ID 768; ORF85-1>:

1 ..VSVGAQASGQ IKILYVKLGQ QVKKGDLIAE INSTSQTNTL NTEKSKLETY 51  QAKLVSAQIA LGSAEKKYKR QAALWKENAT SKEDLESAQD AFAAAKANVA 101  ELKALIRQSK ISINTAESEL GYTRITATMD GTVVAILVEE GQTVNAAQST 151  PTIVQLANLD MMLNKMQIAE GDITKVKAGQ DISFTILSEP DTPIKAKLDS 201  VDPGLTTMSS GGYNSSTDTA SNAVYYYARS FVPNPDGKLA TGMTTQNTVE 251  IDGVKNVLII PSLTVKNRGG KAFVRVLGAD GKAAEREIRT GMRDSMNTEV 301  KSGLKEGDKV VISEITAAEQ QESGERALGG PPRR*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF85 shows 87.8% identity over a 41 aa overlap and 99.3% identity overa 153aa overlap with an ORF (ORF85a) from strain A of N. meningitidis:

The complete length ORF85a nucleotide sequence <SEQ ID 769> is:

1 ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG CGGCGGCAGC 51GGTTTGGGGC GGATGGTCTT ATCTGAAGCC CGAGCCGCAG GCTGCTTATA 101TTACGGAAAC GGTCAGGCGC GGCGACATCA GCCGGACGGT TTCTGCAACA 151GGGGAGATTT CGCCGTCCAA CCTGGTATCG GTCGGCGCGC AGGCATCGGG 201GCAGATTAAG AAACTTTATG TCAAACTCGG GCAACAGGTT AAAAAGGGCG 251ATTTGATTGC GGAAATCAAT TCGACCTCGC AGACCAATAC GCTCAATACG 301GAAAAATCCA AATTGGAAAC GTATCAGGCG AAGCTGGTGT CGGCACAGAT 351TGCATTGGGC AGCGCGGAGA AGAAATATAA GCGTCAGGCG GCGTTGTGGA 401AGGATGATGC GACCGCTAAA GAAGATTTGG AAAGCGCACA GGATGCGCTT 451GCCGCCGCCA AAGCCAATGT TGCCGAGCTG AAGGCTCTAA TCAGACAGAG 501CAAAATTTCC ATCAATACCG CCGAGTCGGA ATTGGGCTAC ACGCGCATTA 551CCGCAACGAT GGACGGCACG GTGGTGGCGA TTCTCGTGGA AGAGGGGCAG 601ACTGTGAACG CGGCGCAGTC TACGCCGACG ATTGTCCAAT TGGCGAATCT 651GGATATGATG TTGAACAAAA TGCAGATTGC CGAGGGCGAT ATTACCAAGG 701TGAAGGCGGG GCAGGATATT TCGTTTACGA TTTTGTCCGA ACCGGATACG 751CCGATTAAGG CGAAGCTCGA CAGCGTCGAC CCCGGGCTGA CCACGATGTC 801GTCGGGCGGC TACAACAGCA GTACGGATAC GGCTTCCAAT GCGGTCTACT 851ATTATGCCCG TTCGTTTGTG CCGAATCCGG ACGGCAAACT CGCCACGGGG 901ATGACGACGC AGAATACGGT TGAAATCGAC GGTGTGAAAA ATGTGCTGAT 951TATTCCGTCG CTGACCGTGA AAAATCGCGG CGGCAGGGCG TTTGTGCGCG 1001TGTTGGGTGC AGACGGCAAG GCGGCGGAAC GCGAAATCCG GACCGGTATG 1051AGAGACAGTA TGAATACCGA AGTAAAAAGC GGGTTGAAAG AGGGGGACAA 1101AGTGGTCATC TCCGAAATAA CCGCCGCCGA GCAGCAGGAA AGCGGCGAAC 1151GCGCCCTAGG CGGCCCGCCG CGCCGATAA

This encodes a protein having amino acid sequence <SEQ ID 770>:

1 MAKMMKWAAV AAVAAAAVWG GWSYLKPEPQ AAYITETVRR GDISRTVSAT 51GEISPSNLVS VGAQASGQIK KLYVKLGQQV KKGDLIAEIN STSQTNTLNT 101EKSKLETYQA KLVSAQIALG SAEKKYKRQA ALWKDDATAK EDLESAQDAL 151AAAKANVAEL KALIRQSKIS INTAESELGY TRITATMDGT VVAILVEEGQ 201TVNAAQSTPT IVQLANLDMM LNKMQIAEGD ITKVKAGQDI SFTILSEPDT 251PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV PNPDGKLATG 301MTTQNTVEID GVKNVLIIPS LTVKNRGGRA FVRVLGADGK AAEREIRTGM 351RDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP RR*

ORF85a and ORF85-1 show 98.2% identity in 334 aa overlap:

FIG. 19D shows plots of hydrophilicity, antigenic index, and AMPHIregions for ORF85a.

Homology with a Predicted ORF from N. gonorrhoeae

ORF85 shows a high degree of identity with a predicted ORF (ORF85ng)from N. gonorrhoeae:

The complete length ORF85ng nucleotide sequence <SEQ ID 771> is:

1 ATGGCAAAAA TGATGAAATG GGCGGCTGTT GCGGCGGTCG CGGCGGCaac 51GGTTTGGGGC GGATGGTCTT ATCTGAAGCC CGAACCGCAG GCTGCTTATA 101TTACGGAaac ggTCAGGCGC GGCGATATCA GCCGGACGGT TTCCGCGACG 151GgcgAGATTT CGCCGTCCAA CCTGGTATCG GTCGGCGCGC AGGCTTCGGG 201GCAGATTAAA AAGCTTTATG TCAAACTCGG GCAACAGGTC AAAAAGGGCG 251ATTTGATTGC GGAAATCAAT TCGACCACGC AGACCAACAC GATCGATATG 301GAAAAATCCA AATTGGAAAC GTATCAGGCG AAGCTGGTGT CGGCACAGAT 351TGCATTGGGC AGCGCGGAGA AGAAATATAA GCGTCAGGCG GCGTTGTGGA 401AGGATGATGC GACCTCTAAA GAAGATTTGG AAAGCGCGCA GGATGCGCTT 451GCCGCCGCCA AAGCCAATGT TGCCGAGTTG AAGGCTTTAA TCAGACAGAG 501CAAAATTTCC ATCAATACCG CCGAGTCGGA TTTGGGCTAC ACGCGCATTA 551CCGCGACGAT GGACGGCACG GTGGTGGCGA TTCCCGTGGA AGAGGGGCAG 601ACTGTGAACG CGGCGCAGTC TACGCCGACG ATTGTCCAAT TGGCGAATCT 651GGATATGATG TTGAACAAAA TGCAGATTGC CGAGGGCGAT ATTACCAAGG 701TGAAGGCGGG GCAGGATATT TCGTTTACGA TTTTGTCCGA ACCGGATACG 751CCGATTAAGG CGAAGCTCGA CAGCGTCGAC CCCGGGCTGA CCACGATGTC 801GTCGGGCGGC TACAACAGCA GTACGGATAC GGCTTCCAAT GCGGTCTATT 851ATTATGCCCG TTCGTTTGTG CCGAATCCGG ACGGCAAACT CGCCACGGGG 901ATGACGACGC AGAATACGGT TGAAATCGAC GGTGTGAAAA ATGTGTTGCT 951TATTCCGTCG CTGACCGTGA AAAATCGCGG CGGCAAGGCG TTCGTACGCG 1001TGTTGGGTGC GGACGGCAAG GCAGTGGAAC GCGAAATCCG GACCGGTATG 1051AAAGACAGTA TGAATACCGA AGTGAAAAGC GGGTTGAAAG AGGGGGACAA 1101AGTGGTCATC TCCGAAATAA CCGCCGCCGA GCAGCAGGAA AGCGGCGAAC 1151GCGCCCTAGG CGGCCCGCCG CGCCGATAA

This encodes a protein having amino acid sequence <SEQ ID 772>:

1 MAKMMKWAAV AAVAAAAVWG GWSYLKPEPQ AAYITEAVRR GDISRTVSAT 51GEISPSNLVS VGAQASGQIK KLYVKLGQQV KKGDLIAEIN STTQTNTIDM 101EKSKLETYQA KLVSAQIALG SAEKKYKRQA ALWKDDATSK EDLESAQDAL 151AAAKANVAEL KALIRQSKIS INTAESDLGY TRITATMDGT VVAIPVEEGQ 201TVNAAQSTPT IVQLANLDMM LNKMQIAEGD ITKVKAGQDI SFTILSEPDT 251PIKAKLDSVD PGLTTMSSGG YNSSTDTASN AVYYYARSFV PNPDGKLATG 301MTTQNTVEID GVKNVLLIPS LTVKNRGGKA FVRVLGADGK AVEREIRTGM 351KDSMNTEVKS GLKEGDKVVI SEITAAEQQE SGERALGGPP RR*

ORF85ng and ORF85-1 show 96.1% identity in 334 aa overlap:

In addition, ORF85ng shows significant homology to an E. coli membranefusion protein:

gi|1787104 (AE000189) o380; 27% identical (27 gaps) to 332 residues frommembrane fusion protein precursor, MTRC_NEIGO SW: P43505 (412 aa)[Escherichia coli] Length = 380 Score = 193 bits (485), Expect = 2e−48Identities = 120/345 (34%), Positives = 182/345 (51%), Gaps =13/345 (3%) Query: 29PQAAYITETVRRGDISRTVSATGEISPSNLVSVGAQASGQIKKLYVKLGQQVKKGDLIAE 88P   Y T  VR GD+ ++V ATG++     V VGAQ SGQ+K L V +G +VKK  L+ Sbjct: 41PVPTYQTLIVRPGDLQQSVLATGKLDALRKVDVGAQVSGQLKTLSVAIGDKVKKDQLLGV 100 Query:89 INSTTQTNTIDMEKSKLETYQAKLVSAQIALGSAEKKYKRQAALWKDDATSKEXXXXXXX 148 I+    N I   ++ L   +A+   A+  L  A   Y RQ  L +  A S++ Sbjct: 101IDPEQAENQIKEVEATLMELRAQRQQAEAELKLARVTYSRQQRLAQTKAVSQQDLDTAAT 160 Query:149 XXXXXXXXXXXXXXXIRQSKISINTAESDLGYTRITATMDGTVVAIPVEEGQTVNAAQST 208               I++++ S++TA+++L YTRI A M G V  I   +GQTV AAQ Sbjct: 161EMAVKQAQIGTIDAQIKRNQASLDTAKTNLDYTRIVAPMAGEVTQITTLQGQTVIAAQQA 220 Query:209 PTIVQLANLDMMLNKMQIAEGDITKVKAGQDISFTILSEPDTPIKAKLDSVDPGLTTMSS 268P I+ LA++  ML K Q++E D+  +K GQ   FT+L +P T  + ++  V P Sbjct: 221PNILTLADMSAMLVKAQVSEADVIHLKPGQKAWFTVLGDPLTRYEGQIKDVLP------- 273 Query:269 GGYNSSTDTASNAVYYYARSFVPNPDGKLATGMTTQNTVEIDGVKNVLLIPSLTVKNRGG 328     + +  ++A++YYAR  VPNP+G L   MT Q  +++  VKNVL IP   + +  G Sbjct: 274-----TPEKVNDAIFYYARFEVPNPNGLLRLDMTAQVHIQLTDVKNVLTIPLSALGDPVG 328 Query:329 KAFVRV-LGADGKAVEREIRTGMKDSMNTEVKSGLKEGDKVVISE 372     +V L  +G+ ERE+  G ++  + E+  GL+ GD+VVI E Sbjct: 329DNRYKVKLLRNGETREREVTIGARNDTDVEIVKGLEAGDEVVIGE 373

Based on this analysis, it was predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF85-1 (40.4 kDa) was cloned in the pGex vectors and expressed in E.coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 19A shows the results ofaffinity purification of the GST-fusion protein. Purified GST-fusionprotein was used to immunise mice, whose sera were used for Western blot(FIG. 19B), FACS analysis (FIG. 19C), and ELISA (positive result). Theseexperiments confirm that ORF85-1 is a surface-exposed protein, and thatit is a useful immunogen.

Example 92

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 773>:

1 ..ATTCCCGCCA CGATGACATT TGAACGCAGC GGCAATGCTT ACAAAATCGT 51  TTCGACGATT AAAGTGCCGC TATACAATAT CCGTTTCGAG TCCGGCGGTA 101  CGGTTGTCGG CAATACCCTG CACCCTACCT ACTATAGAGA CATACGCAGG 151  GGCAAACTGT ATGCGGAAgc CAAATTCGCC GACgGcAGCG TAACTTACGG 201  CAAAGCGGGC GAGAGCAAAA CCGAGCAAAG CCCCAAGGCT ATGGATTTGT 251  TCACGCTTGC CTGGCAGTTG GCGGCAAATG ACGCGAAACT CCCCCCGGGG 301  CTGAAAATCA CCAACGGCAA AAAACTTTAT TCCGTCGGCG GTTTGAATAA 351  GGCGGGTACA GGAAAATACA GCATAGGCGG CGTGGAAACC GAAGTCGTCA 401  AATATCGGGT GCGGCGCGGC GACGATGCGG TAATGTATTT cTTCGCACCG 451  TCCCTGAACA ATATTCCGGC ACAAATCGGC TATACCGACG ACGGCAAAAC 501  CTATACGCTG AAACTCAAAT CGGTGCAGAT CAACGGCCAG GCAGCCAAAC 551   CGTAA

This corresponds to the amino acid sequence <SEQ ID 774; ORF120>:

1 ..IPATMTFERS GNAYKIVSTI KVPLYNIRFE SGGTVVGNTL HPTYYRDIRR 51  GKLYAEAKFA DGSVTYGKAG ESKTEQSPKA MDLFTLAWQL AANDAKLPPG 101  LKITNGKKLY SVGGLNKAGT GKYSIGGVET EVVKYRVRRG DDAVMYFFAP 151  SLNNIPAQIG YTDDGKTYTL KLKSVQINGQ AAKP*

Further work revealed the complete nucleotide sequence <SEQ ID 775>:

1 ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT TGTCCGCCGC 51 CCTGCCGTGCGCGTATGCGG CAGGGCTGCC CCAATCCGCC GTGCTGCACT 101 ATTCCGGCAG CTACGGCATTCCCGCCACGA TGACATTTGA ACGCAGCGGC 151 AATGCTTACA AAATCGTTTC GACGATTAAAGTGCCGCTAT ACAATATCCG 201 TTTCGAGTCC GGCGGTACGG TTGTCGGCAA TACCCTGCACCCTACCTACT 251 ATAGAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA ATTCGCCGAC301 GGCAGCGTAA CTTACGGCAA AGCGGGCGAG AGCAAAACCG AGCAAAGCCC 351CAAGGCTATG GATTTGTTCA CGCTTGCCTG GCAGTTGGCG GCAAATGACG 401 CGAAACTCCCCCCGGGGCTG AAAATCACCA ACGGCAAAAA ACTTTATTCC 451 GTCGGCGGTT TGAATAAGGCGGGTACAGGA AAATACAGCA TAGGCGGCGT 501 GGAAACCGAA GTCGTCAAAT ATCGGGTGCGGCGCGGCGAC GATGCGGTAA 551 TGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACAAATCGGCTAT 601 ACCGACGACG GCAAAACCTA TACGCTGAAA CTCAAATCGG TGCAGATCAA651 CGGCCAGGCA GCCAAACCGT AA

This corresponds to the amino acid sequence <SEQ ID 776; ORF120-1>:

1 MMKTFKNIFS AAILSAALPC AYAAGLPQSA VLHYSGSYGI PATMTFERSG 51 NAYKIVSTIKVPLYNIRFES GGTVVGNTLH PTYYRDIRRG KLYAEAKFAD 101 GSVTYGKAGE SKTEQSPKAMDLFTLAWQLA ANDAKLPPGL KITNGKKLYS 151 VGGLNKAGTG KYSIGGVETE VVKYRVRRGDDAVMYFFAPS LNNIPAQIGY 201 TDDGKTYTLK LKSVQINGQA AKP*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF120 shows 92.4% identity over a 184aa overlap with an ORF (ORF120a)from strain A of N. meningitidis:

The complete length ORF120a nucleotide sequence <SEQ ID 777> is:

1 ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT TGTCCGCCGC 51 CCTGCCGTGCGCGTATGCGG CAGGGCTGCC CNAATCCGCC GTGCTGCACT 101 ATTCCGGCAG CTACGGCATTCCCGCCACNA NNANNTNNGN ACNNNGNGNC 151 AATGCTTNCA AAATCGTTTC GACGATTAAAGTGCCGCTAT ACAATATCCG 201 TTTCGAGTCC GGCGGTACGG TTGTCGGCAA TACCCTGCACCCTACCTACT 251 ATAGAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA ATTCGCCGAC301 GGCAGCGTAA CCTACGGCAA AGCGGNNNNN ANCNNNNNNG NGCAAAGCCC 351CAAGGCTATG GATTTGTTCA CGCTTGCNTG GCAGTTGGCG GCAAATGACG 401 CGAAACTCCCCCCGGGGCTG AAAATCACCA ACGGCAAAAA ACTTTATTCC 451 GTCGGCGGTT TGAATAAGGCGGGTACAGGA AAATACAGCA TAGGCGGCGT 501 GGAAACCGAA GTCGTCAAAT ATCGGGTGCGGCGCGGCGAC GATGCGGTAA 551 TGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACAAATCGGCTAT 601 ACCGACGACG GCAAAACCTA TACGCTGAAA CTCAAATCGG TGCAGATCAA651 CGGCCAGGCA GCCAAACCGT AA

This encodes a protein having amino acid sequence <SEQ ID 778>:

1 MMKTFKNIFS AAILSAALPC AYAAGLPXSA VLHYSGSYGI PATXXXXXXX 51 NAXKIVSTIKVPLYNIRFES GGTVVGNTLH PTYYRDIRRG KLYAEAKFAD 101 GSVTYGKAXX XXXXQSPKAMDLFTLAWQLA ANDAKLPPGL KITNGKKLYS 151 VGGLNKAGTG KYSIGGVETE VVKYRVRRGDDAVMYFFAPS LNNIPAQIGY 201 TDDGKTYTLK LKSVQINGQA AKP*

ORF120a and ORF120-1 show 93.3% identity in 223 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF120 shows 97.8% identity over 184 aa overlap with a predicted ORF(ORF120ng) from N. gonorrhoeae:

The complete length ORF120ng nucleotide sequence <SEQ ID 779> is:

1 ATGATGAAGA CTTTTAAAAA TATATTTTCC GCCGCCATTT TGTCCGCCGC 51 CCTGCCGTGCGCGTATGCGG CAAGGCTACC CCAATCCGCC GTGCTGCACT 101 ATTCCGGCAG CTACGGCATTCCCGCCACGA TGACATTTGA ACGCAGCGGC 151 AATGCTTACA AAATCGTTTC GACGATTAAAGTGCCGCTAT ACAATATCCG 201 TTTCGAATCC GGCGGTACGG TTGTCGGCAA TACCCTGCACCCTGCCTACT 251 ATAAAGACAT ACGCAGGGGC AAACTGTATG CGGAAGCCAA ATTCGCCGAC301 GGCAGCGTAA CCTACGGCAA AGCGGGCGAG AGCAAAACCG AGCAAAGCCC 351CAAGGCTATG GATTTGTTCA CGCTTGCCTG GCAGTTGGCG GCAAATGACG 401 CGAAACTCCCCCCGGGTCTG AAAATCACCA ACGGCAAAAA ACTTTATTCC 451 GTCGGCGGCC TGAATAAGGCGGGTACGGGA AAATACAGCA TaggCGGCGT 501 GGAAACCGAA GTCGTCAAAT ATCGGGTGCGGCGCGGCGAC GATACGGTAA 551 CGTATTTCTT CGCACCGTCC CTGAACAATA TTCCGGCACAAATCGGCTAT 601 ACCGACGACG GCAAAACCTA TACGCTGAAG CTCAAATCGG TGCAGATCAA651 CGGACAGGCC GCCAAACCGT AA

This encodes a protein having amino acid sequence <SEQ ID 780>:

1 MMKTFKNIFS AAILSAALPC AYAARLPQSA VLHYSGSYGI PATMTFERSG 51 NAYKIVSTIKVPLYNIRFES GGTVVGNTLH PAYYKDIRRG KLYAEAKFAD 101 GSVTYGKAGE SKTEQSPKAMDLFTLAWQLA ANDAKLPPGL KITNGKKLYS 151 VGGLNKAGTG KYSIGGVETE VVKYRVRRGDDTVTYFFAPS LNNIPAQIGY 201 TDDGKTYTLK LKSVQINGQA AKP*

In comparison with ORF120-1, ORF120ng shows 97.8% identity in 223 aaoverlap:

This analysis, including the presence of a putative leader sequence inthe gonococcal protein suggests that the proteins from N. meningitidisand N. gonorrhoeae, and, their epitopes, could be useful antigens forvaccines or diagnostics, or for raising antibodies.

Example 93

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 781>:

1 ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG GTGCCGGTGC 51 .GCGTTTGCCGCCTTGGTCT GGCTGGTTTT CGCGCTCGGC GATACTTTGA 101 CTCCGTTTGC GGTTGCGGCGGTGCTGGCGT ATGTATTGGA CCCTTTGGTC 151 GAATGGTTGC AGAAAAAGGG TTTGAACCGTGCATCCGCTT CGATGTCTGT 201 GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTGATTATCGTCC 251 CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT GCCCCAATTA301 ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA ATACAATCGG 351CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG CTTCAGGCGC 401 ATACGGGAGAGTTGAGCAAC GCGCTTAAGG CGTGGTTTCC CGTTTTGATG 451 AGGCAGGGCG GCAATATT..

This corresponds to the amino acid sequence <SEQ ID 782; ORF121>:

1 MYRRKGRGIK PWMGAGXAFA ALVWLVFALG DTLTPFAVAA VLAYVLDPLV 51 EWLQKKGLNRASASMSVMVF SLILLLALLL IIVPMLVGQF NNLASRLPQL 101 IGFMQNTLLP WLKNTIGGYVEIDQASIIAW LQAHTGELSN ALKAWFPVLM 151 RQGGNI..

Further work revealed the complete nucleotide sequence <SEQ ID 783>:

1 ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG GTGCCGGTGC 51 GGCGTTTGCCGCCTTGGTCT GGCTGGTTTT CGCGCTCGGC GATACTTTGA 101 CTCCGTTTGC GGTTGCGGCGGTGCTGGCGT ATGTATTGGA CCCTTTGGTC 151 GAATGGTTGC AGAAAAAGGG TTTGAACCGTGCATCCGCTT CGATGTCTGT 201 GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTGATTATCGTCC 251 CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT GCCCCAATTA301 ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA ATACAATCGG 351CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG CTTCAGGCGC 401 ATACGGGAGAGTTGAGCAAC GCGCTTAAGG CGTGGTTTCC CGTTTTGATG 451 AGGCAGGGCG GCAATATTGTCAGCAGTATC GGCAACCTGC TGCTGCTTCC 501 CTTGCTGCTT TACTATTTCC TGCTGGATTGGCAGCGGTGG TCGTGCGGCA 551 TTGCCAAACT GGTTCCGAgG CGTTTTGCCG GTGCTTATACGCGCATTACA 601 GGCAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGGC AGCTTCTGGT651 AATGCTGATT ATGGGCTTGG TTTACGGTTT GGGATTGGTG CTGGTCGGGC 701TGGATTCGGG GTTTGCCATC GGTATGCTTG CCGGTATTTT GGTGTTTGTC 751 CCTTATCTCGGGGCGTTTAC GGGATTGCTG CTTGCCACCG TCGCCGCCTT 801 GCTCCAGTTC GGTTCGTGGAACGGCATCCT ATCGGTTTGG GCGGTTTTTG 851 CCGTAGGACA GTTTCTCGAA AGTTTTTTCATTACGCCGAA AATCGTGGGA 901 GACCGTATCG GGCTGTCGCC GTTTTGGGTT ATCTTTTCGCTGATGGCGTT 951 CGGGCAGCTG ATGGGCTTTG TCGGAATGTT GGCGGGATTG CCTTTGGCCG1001 CCGTAACCTT GGTCTTGCTT CGCGAGGGCG TGCAGAAATA TTTTGCCGGC 1051AGTTTTTACC GGGGCAGGTA G

This corresponds to the amino acid sequence <SEQ ID 784; ORF121-1>:

1 MYRRKGRGIK PWMGAGAAFA ALVWLVFALG DTLTPFAVAA VLAYVLDPLV 51 EWLQKKGLNRASASMSVMVF SLILLLALLL IIVPMLVGQF NNLASRLPQL 101 IGFMQNTLLP WLKNTIGGYVEIDQASIIAW LQAHTGELSN ALKAWFPVLM 151 RQGGNIVSSI GNLLLLPLLL YYFLLDWQRWSCGIAKLVPR RFAGAYTRIT 201 GNLNEVLGEF LRGQLLVMLI MGLVYGLGLV LVGLDSGFAIGMLAGILVFV 251 PYLGAFTGLL LATVAALLQF GSWNGILSVW AVFAVGQFLE SFFITPKIVG301 DRIGLSPFWV IFSLMAFGQL MGFVGMLAGL PLAAVTLVLL REGVQKYFAG 351 SFYRGR*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF121 shows 98.7% identity over a 156aa overlap with an ORF (ORF121a)from strain A of N. meningitidis:

The complete length ORF121a nucleotide sequence <SEQ ID 785> is:

1 ATGTATCGGA GGAAAGGGCG GGGCATCAAG CCGTGGATGG ATGCCGGTGC 51 GGCGTTTGCCGCCTTGGTCT GGCTGGTTTT CGCGCTCGGC GATACTTTGA 101 CTCCGTTTGC GGTTGCGGCGGTGCTGGCGT ATGTATTGGA CCCTTTGGTC 151 GAATGGTTGC AGAAAAAGGG TTTGAACCGTGCATCCGCTT CGATGTCTGT 201 GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTGATTATTGTCC 251 CTATGCTGGT CGGGCAGTTC AACAATTTGG CATCGCGCCT GCCCCAATTA301 ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA ATACAATCGG 351CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG CTTCAGGCGC 401 ATACGGGCGAGTTGAGCAAC GCGCTTAAGG CGTGGTTTCC CGTTTTGATG 451 AGGCAGGGCG GCAATATTGTCAGCAGTATC GGCAACCTGC TGCTGCTTCC 501 CTTGCTGCTT TACTATTTCC TGCTGGATTGGCAGCGGTGG TCGTGCGGCA 551 TTGCCAAACT GGTTCCGAGG CGTTTTGCCG GTGCTTATACGCGCATTACA 601 GGCAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGGC AGCTTCTGGT651 GATGCTGATT ATGGGTTTGG TTTACGGCTT GGGGTTGGTG CTGGTCGGGC 701TGGATTCGGG GTTTGCAATC GGTATGGTTG CCGGTATTTT GGTTTTTGTT 751 CCCTATTTGGGCGCGTTTAC AGGACTGCTG CTGGCAACCG TCGCCGCCTT 801 GCTCCAGTTC GGTTCGTGGAACGGCATCTT GGCTGTTTGG GCGGTTTTTG 851 CCGTAGGACA GTTTCTCGAA AGTTTTTTCATTACGCCGAA AATCGTGGGA 901 GACCGTATCG GCCTGTCGCC GTTTTGGGTT ATCTTTTCGCTGATGGCGTT 951 CGGGCAGCTG ATGGGCTTTG TCGGAATGTT GGCCGGATTG CCTTTGGCCG1001 CCGTAACCTT GGTCTTGCTT CGCGAGGGCG TGCAGAAATA TTTTGCCGGC 1051AGTTTTTACC GGGGCAGGTA G

This encodes a protein having amino acid sequence <SEQ ID 786>:

1 MYRRKGRGIK PWMDAGAAFA ALVWLVFALG DTLTPFAVAA VLAYVLDPLV 51 EWLQKKGLNRASASMSVMVF SLILLLALLL IIVPMLVGQF NNLASRLPQL 101 IGFMQNTLLP WLKNTIGGYVEIDQASIIAW LQAHTGELSN ALKAWFPVLM 151 RQGGNIVSSI GNLLLLPLLL YYFLLDWQRWSCGIAKLVPR RFAGAYTRIT 201 GNLNEVLGEF LRGQLLVMLI MGLVYGLGLV LVGLDSGFAIGMVAGILVFV 251 PYLGAFTGLL LATVAALLQF GSWNGILAVW AVFAVGQFLE SFFITPKIVG301 DRIGLSPFWV IFSLMAFGQL MGFVGMLAGL PLAAVTLVLL REGVQKYFAG 351 SFYRGR*

ORF121a and ORF121-1 show 99.2% identity in 356 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF121 shows 97.4% identity over a 156 aa overlap with a predicted ORF(ORF121ng) from N. gonorrhoeae:

An ORF121ng nucleotide sequence <SEQ ID 787> was predicted to encode aprotein having amino acid sequence <SEQ ID 788>:

1 MYRRKGRGIK PWMGAGAAFA ALVWLVYALG DTLTPFAVAA VLAYVLDPLV 51 EWLQKKGLNRASASMSVMVF SLILLLALLL IIVPMLVGQF NNLASRLPQL 101 IGFMQNTLLP WLKNTIGGYVEIDQASIIAW FQAHTGELSN ALKAWFPVLM 151 KQGGNIVSTI GNLLLPPLLL YYFLLDWHRWSCGIPKLVPR RFAGAYTRIT 201 GNLNKVWGKF LRGQLLGETE RGAVVCRVGR ECWEGGGARSRPSDDGWPRW 251 GGG*

Further work revealed the following gonoccocal DNA sequence <SEQ ID789>:

1 ATGTATCGGA GAAAAGGACG GGGCATCAAG CCGTGGATGG GTGCCGGCGC 51 GGCGTTTGCCGCCTTGGTCT GGCTGGTTTA CGCGCTCGGC GATACTTTGA 101 CTCCGTTTGC GGTTGCGGCGGTGCTGGCGT ATGTGTTGGA CCCTTTGGTC 151 GAATGGTTGC AGAAAAAGGG TTTGAACCGTGCATCCGCTT CGATGTCTGT 201 GATGGTGTTT TCCTTGATTT TGTTGTTGGC ATTATTGTTGATTATTGTCC 251 CTATGCTGGT CGGGCAGTTC AATAATTTGG CATCTCGCCT GCCCCAATTA301 ATCGGTTTTA TGCAGAACAC GCTGCTGCCG TGGTTGAAAA ATACAATCGG 351CGGATATGTG GAAATCGATC AGGCATCTAT TATTGCGTGG TTTCAGGCGC 401 ATACGGGCGAGTTGAGCAAC GCGCTTAAGG CGTGGTTTCC CGTTTTGATG 451 AAACAGGGCG GCAATATTGTCAGCAGTATC GGCAACCTGC TGCTGCCGCC 501 CTTGCTGCTT TACTATTTCC TGCTGGATTGGCAGCGGTGG TCGTGCGGCA 551 TCGCCAAACT GGTTCCGAGG CGTTTTGCCG GTGCTTATACGCGCATTACG 601 GGTAATTTGA ACGAGGTATT GGGCGAATTT TTGCGCGGTC AGCTTCTGGT651 GATGCTGATT ATGGGCTTGG TTTACGGTTT GGGATTGATG CTAGTCGGAC 701TGGATTCGGG ATTTGCCATC GGTATGGTTG CCGGTATTTT GGTGTTTGTC 751 CCCTATTTGGGTGCGTTTAC GGGATTGCTG CTTGCCACTG TTGCAGCCTT 801 GCTCCAGTTC GGTTCGTGGAACGGAATCTT GGCTGTTTGG GCGGTTTTTG 851 CCGTCGGTCA GTTTCTCGAA AGTTTTTTCATTACGCCGAA AATTGTAGGA 901 GACCGTATCG GCCTGTCGCC GTTTTGGGTT ATCTTTTCGCTGATGGCGTT 951 CGGAGAGCTG ATGGGCTTTG TCGGAATGTT GGCCGGATTG CCTTTGGCCG1001 CCGTAACCTT GGTCTTGCTT CGCGAGGGCG CGCAGAAATA TTTTGCCGGC 1051AGTTTTTACC GGGGCAGGTA G

This corresponds to the amino acid sequence <SEQ ID 790; ORF121ng-1>:

1 MYRRKGRGIK PWMGAGAAFA ALVWLVYALG DTLTPFAVAA VLAYVLDPLV 51 EWLQKKGLNRASASMSVMVF SLILLLALLL IIVPMLVGQF NNLASRLPQL 101 IGFMQNTLLP WLKNTIGGYVEIDQASIIAW FQAHTGELSN ALKAWFPVLM 151 KQGGNIVSSI GNLLLPPLLL YYFLLDWQRWSCGIAKLVPR RFAGAYTRIT 201 GNLNEVLGEF LRGQLLVMLI MGLVYGLGLM LVGLDSGFAIGMVAGILVFV 251 PYLGAFTGLL LATVAALLQF GSWNGILAVW AVFAVGQFLE SFFITPKIVG301 DRIGLSPFWV IFSLMAFGEL MGFVGMLAGL PLAAVTLVLL REGAQKYFAG 351 SFYRGR*

ORF121ng-1 and ORF121-1 show 97.5% identity in 356 aa overlap:

In addition, ORF121ng-1 shows homology to a permease from H. influenzae:

sp|P43969|PERM_HAEIN PUTATIVE PERMEASE PERM HOMOLOG Length = 349 Score= 69.9 bits (168), Expect = 2e−11 Identities = 67/317 (21%), Positives= 120/317 (37%), Gaps = 7/317 (2%) Query: 26VYALGDTLTPFAVAAVLAYVLDPLVEWL-QKKGLNRASASMSVMVFSXXXXXXXXXXXVP 84 +Y  GD+ P  +A VL+Y+L+  + +L Q     R  A++ +               VP Sbjct: 32IYFFGDLIAPLLIALVLSYLLEIPINFLNQYLKCPRMLATILIFGSFIGLAAVFFLVLVP 91 Query:85 MLVGQFNNLASRLPQLIGFMQNTLLPWLKNTIGGYVE-IDQASIIAWFQAHTGELSNALK 143ML  Q  +L S LP +     N    WL N    Y E ID + + + F +   ++    + Sbjct: 92MLWNQTISLLSDLPAMF----NKSNEWLLNLPKNYPELIDYSMVDSIFNSVREKILGFGE 147 Query:144 AWFPVLMKQGGNIVSSIGNXXXXXXXXXXXXXDWQRWSCGIAKLVPRRFAGAYTRITGNL 203+   + +    N+VS                 D      G+++ +P+    A+ R    + Sbjct: 148SAVKLSLASIMNLVSLGIYAFLVPLMMFFMLKDKSELLQGVSRFLPKNRNLAFXRWK-EM 206 Query:204 NEVLGEFLRGQXXXXXXXXXXXXXXXXXXXXDSGFAIGMVAGILVFVPYXXXXXXXXXXX 263 + +  ++ G+                    +    +    G+ V VPY Sbjct: 207QQQISNYINGKLLEILIVTLITYIIFLIFGLNYPLLLAFAVGLSVLVPYIGAVIVTIPVA 266 Query:264 XXXXXQFGSWNGILAVWAVFAVGQFLESFFITPKIVGDRIGLSPFWVIFSLMAFGELMGF 323     QFG       +   FAV Q L+   + P +  + + L P  +I S++ FG L GF Sbjct: 267LVALFQFGISPTFWYIIIAFAVSQLLDGNLLVPYLFSEAVNLHPLIIIISVLIFGGLWGF 326 Query:324 VGMLAGLPLAAVTLVLL 340  G+   +PLA +   ++ Sbjct: 327 WGVFFAIPLATLVKAVI343

Based on this analysis, including the presence of a putative leadersequence and transmembrane domains in the two proteins, it is predictedthat the proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 94

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 791>:

1 ..ACTGCTTTTT CGGCGGCGCT GCGCTTGAGT CCATCATGAC TCGTCATATT 51  TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATC TTAACATTTT 101  TTTGCACGTC CTGCCCGCCG CGTTCAAATG CGTACCAGCA ATACCGCCGC 151  CTGCGCCTCT ATGCCTTCCA TCCGCCCGAG ATAGCCGAGT TTTTCGTTGG 201  TTTTGCCTTT GATGTTGACG CACGAAATGT CTATGCCCAA ATCGGCGGCG 251  ATGTTGGCAC GCATTTGCGG AATGTGCGGC GCGAGTGTGG GTTTCTGTGC 301  AATCACGGTC GTATCGACAT TGACCGCCTG CCAACCCTGC GCCTGAACGC 351  TTTGATACGC CGCACGCAAA AGGACGCGGC TGTCCGCATC TTTGAACTCT 401  GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAAC CTGCCGCACC 451  GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA TCGGAGTGTC 501  CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG TATCAG..

This corresponds to the amino acid sequence <SEQ ID 792; ORF122>:

1 ..TAFSAALRLS PSXLVIFLSF GKPYQQTAAI LTFFCTSCPP RSNAYQQYRR 51  LRLYAFHPPE IAEFFVGFAF DVDARNVYAQ IGGDVGTHLR NVRRECGFLC 101  NHGRIDIDRL PTLRLNALIR RTQKDAAVRI FELCGGVGEM AADIAQTCRT 151  EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAK YQ..

Further work revealed the complete nucleotide sequence <SEQ ID 793>:

1 ATATCGTACT GGGCAAGCAG TTCGCCGGAT TTTTTGGAAG TAGATACCGC 51 GCCTTTGATTTTTTTGCCGC TCTTACCCAA GGCTTCGATG AAAAAGTTGA 101 TGGTCGAGCC GGTACCGATGCCGATATATT CATTTTCGGG TACGAATTCG 151 ACTGCTTTTT CGGCGGCGAT GCGCTTGAGTTCGTCTTGTG TCGTCATATT 201 TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATCTTAACATTTT 251 TTTGCACGTC CTGCCCGCCG CGTTCAAATG CGTACCAGCA ATACCGCCGC301 CTGCGCCTCT ATGCCTTCCA TCCGCCCGAG ATAGCCGAGT TTTTCGTTGG 351TTTTGCCTTT GATGTTGACG CACGAAATGT CTATGCCCAA ATCGGCGGCG 401 ATGTTGGCACGCATTTGCGG AATGTGCGGC GCGAGTTTGG GTTTCTGTGC 451 AATCACGGTC GTATCGACATTGACCGCCTG CCAACCCTGC GCCTGAACGC 501 TTTGATACGC CGCACGCAAA AGGACGCGGCTGTCCGCATC TTTGAACTCT 551 GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAACCTGCCGCACC 601 GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA TCGGAGTGTC651 CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG TATCAGCTTT 701CTGCCTTCGG TCAGTTGGTG GACATCGTAG CCCTGTCCGA TACGGATGTT 751 CGTCATCGTTTGTGTTCCTG A

This corresponds to the amino acid sequence <SEQ ID 794; ORF122-1>:

1 ISYWASSSPD FLEVDTAPLI FLPLLPKASM KKLMVEPVPM PIYSFSGTNS 51TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFCTSCPP RSNAYQQYRR 101 LRLYAFHPPEIAEFFVGFAF DVDARNVYAQ IGGDVGTHLR NVRREFGFLC 151 NHGRIDIDRL PTLRLNALIRRTQKDAAVRI FELCGGVGEM AADIAQTCRT 201 EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAKYQLSAFGQLV DIVALSDTDV 251 RHALCS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF122 shows 94.0% identity over a 182aa overlap with an ORF (ORF122a)from strain A of N. meningitidis:

The complete length ORF122a nucleotide sequence <SEQ ID 795> is:

1 ATATCATATT GGGCAAGCAG TTCACTGGAT TTTTTGGAAG TAGATACCGC 51 GCCTTTGATTTTTTTGCCGC TCTTACCCAA GGCTTCGATG AAAAAGTTGA 101 TGGTCGAACC GGTACCGATGCCGATGTATT CGTTTTCGGG TACGAATTCG 151 ACTGCNTTTT CGGCGGCGAT GCGCTTGAGTTCGTCTTGTG TCGTCATATT 201 TTTGTCCTTT GGGAAACCGT ATCAACAAAC AGCCGCCATCTTAACATTTT 251 TTNNNACGTC CTGCCCGCCG CGTTCAAATC CTTACCAGCA ATACCGCCGC301 CTGCGACTCT ATGCCTTCCA TGCGCCCGAG ATAACCGAGT TTTTCGTTGG 351TTTTGCCTTT GANGTTGACG CACGAAATGT CTATGCCCAA ATCGGCGGCG 401 ATGTTGGCACGCATTTGCGG AATATGCGGC GCGAGTTTGG GTTTCTGTGC 451 AATCACGGTC GTATCGACATTGACCGCCTG CCAACCCTGC GCCTGAACGC 501 TTTGATACGC CGCACGCAAA AGGACGCGGCTGTCCGCATC TTTGAACTCT 551 GCGGCGGTGT CGGGGAAATG GCTGCCGATA TCGCCCAAACCTGCCGCACC 601 GAGCAGCGCG TCGGTAACGG CGTGCAGCAG CGCATCGGCA TCGGAGTGTC651 CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG TATCAGCTTT 701CTGCCTTCGG TCAGTTGGTG GACATCGTAG CCCTGTCCGA TACGGATGTT 751 CGTCATCGTTTGTGTTCCTG A

This encodes a protein having amino acid sequence <SEQ ID 796>:

1 ISYWASSSLD FLEVDTAPLI FLPLLPKASM KKLMVEPVPM PMYSFSGTNS 51TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFXTSCPP RSNPYQQYRR 101 LRLYAFHAPEITEFFVGFAF XVDARNVYAQ IGGDVGTHLR NMRREFGFLC 151 NHGRIDIDRL PTLRLNALIRRTQKDAAVRI FELCGGVGEM AADIAQTCRT 201 EQRVGNGVQQ RIGIGVSEQP FFKWDFNSAKYQLSAFGQLV DIVALSDTDV 251 RHRLCS*

ORF122a and ORF122-1 show 96.9% identity in 256 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF122 shows 89.6% identity over a 182 aa overlap with a predicted ORF(ORF122ng) from N. gonorrhoeae:

The complete length ORF122ng nucleotide sequence <SEQ ID 797> is:

1 ATGTCGTACC GGGCAAGCAG TTCGCCGGAT TTTTTGGAGG TTGAAACCGC 51 GCCTTTGATTTTTTTACCGC TTTTGCCCAA GGCTTCGATG AAGAAATTGa 101 tgGTCGAACC GgtaCCGATGCCGATGTATT CGTTTTCGGG TACGAATTCG 151 ACTGCTTTTT CGGCGGCGAT GCGCttgAgtTCgtcttgcg TcgTCATATT 201 TTTAtccttt gGGAAaccct atcaAcaAAc agccgccatCTTAACATTTT 251 TTTGCACGtc ctggccgccg cgttcaAATc cgtaccaGca ataccgccgc301 ctgcgcctCT AtgcCTTCCA TCCGCCCGAG ATAGCCGAGT TTTTCGTTGG 351TTTTGCCTTT GATatTGACG CACGAAATAT CGatacCCAa atcggcgGCG 401 ATGTTGGCACGCATTTGCGG AATGTGCGGT GCGAGTTTGG GTTTCTGTGC 451 AATCACGGTC GTATCGACATTGACCACCTG CCAACCCTGC GCCTGAACGC 501 TTTGATACGC CGCACGCAAA AGGACGCGGCTGTCCGCATC TTTGAACTCT 551 GCGGCGGTGT CGGGAAAATG GCTGCCGATG TCGCCCAAACCTGCCGCACC 601 GAGCAGCgcg tcggtaaCGG CGTGCAGCAG cgcgTcgGCA TCCGAATGCC651 CGAGCAGCCC TTTTTCAAAT GGGATTTCAA CTCCGCCAAG TATCAGCTTT 701CTGCCTTCGG TCAATTGGTG GACATCGTAG CCCTGTCCGA TACGGATATT 751 CGTCATCGTTTGTGTTCCTG A

This encodes a protein having amino acid sequence <SEQ ID 798>:

1 MSYRASSSPD FLEVETAPLI FLPLLPKASM KKLMVEPVPM PMYSFSGTNS 51TAFSAAMRLS SSCVVIFLSF GKPYQQTAAI LTFFCTSWPP RSNPYQQYRR 101 LRLYAFHPPEIAEFFVGFAF DIDARNIDTQ IGGDVGTHLR NVRCEFGFLC 151 NHGRIDIDHL PTLRLNALIRRTQKDAAVRI FELCGGVGKM AADVAQTCRT 201 EQRVGNGVQQ RVGIRMPEQP FFKWDFNSAKYQLSAFGQLV DIVALSDTDI 251 RHRLCS*

ORF122ng and ORF122-1 show 92.6% identity in 256 aa overlap:

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 95

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 799>:

1 ..GCCGGCGCGA GTGCGAACAA CATTTCCGCG CGTTTTGCGG AAACACCCGT 51  CGCTGTCAGC GTTACCCTGA TCGGCACGGT ACTTGCCGTC ATGCTGCCCG 101  TTACCGAATA TGAAAACTTC CTGCTGCTTA TCGGCTCGGT ATTTGCGCCG 151  ATGGGGCGGA TTTTGATTGC CGACTTTTTC GTCTTGAAAC GGCGTGA

This corresponds to the amino acid sequence <SEQ ID 800; ORF125>:

1 ..AGASANNISA RFAETPVAVS VTLIGTVLAV MLPVTEYENF LLLIGSVFAP 51  MGGFDCRLFR LETA*

Further work revealed the complete nucleotide sequence <SEQ ID 801>:

1 ATGTCGGGCA ATGCCTCCTC TCCTTCATCT TCCTCCGCCA TCGGGCTGAT 51 TTGGTTCGGCGCGGCGGTAT CGATTGCCGA AATCAGCACG GGTACGCTGC 101 TTGCGCCTTT GGGCTGGCAGCGCGGTCTGG CGGCTCTACT TTTGGGTCAT 151 GCCGTCGGCG GCGCGCTGTT TTTTGCGGCGGCGTATATCG GCGCACTGAC 201 CGGACGCAGC TCGATGGAAA GCGTGCGCCT GTCGTTCGGCAAACGCGGTT 251 CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG CTGGACGGCG301 GTGATGATTT ACGCCGGCGC AACGGTCAGC TCCGCTTTGG GCAAAGTGTT 351GTGGGACGGC GAATCTTTTG TCTGGTGGGC ATTGGCAAAC GGCGCGCTGA 401 TTGTGCTGTGGCTGGTTTTC GGCGCACGCA AAACAGGCGG GCTGAAAACC 451 GTTTCGATGC TGCTGATGCTGTTGGCGGTT CTGTGGCTGA GTGCCGAAGT 501 CTTTTCCACG GCAGGCAGCA CCGCCGCACAGGTTTCAGAC GGCATGAGTT 551 TCGGAACGGC AGTCGAGCTG TCCGCCGTGA TGCCGCTTTCCTGGCTGCCG 601 CTTGCCGCCG ACTACACGCG CCACGCGCGC CGCCCGTTTG CGGCAACCCT651 GACGGCAACG CTCGCCTACA CGCTGACCGG CTGCTGGATG TATGCCTTGG 701GTTTGGCAGC GGCGTTGTTC ACCGGAGAAA CCGACGTGGC AAAAATCCTG 751 CTGGGCGCAGGTTTGGGTGC GGCAGGCATT TTGGCGGTCG TCCTCTCCAC 801 CGTTACCACA ACGTTTCTCGATGCCTATTC CGCCGGCGCG AGTGCGAACA 851 ACATTTCCGC GCGTTTTGCG GAAACACCCGTCGCTGTCGG CGTTACCCTG 901 ATCGGCACGG TACTTGCCGT CATGCTGCCC GTTACCGAATATGAAAACTT 951 CCTGCTGCTT ATCGGCTCGG TATTTGCGCC GATGGCGGCG GTTTTGATTG1001 CCGACTTTTT CGTCTTGAAA CGGCGTGAGG AGATTGAAGG CTTTGACTTT 1051GCCGGACTGG TTCTGTGGCT TGCGGGCTTC ATCCTCTACC GCTTCCTGCT 1101 CTCGTCCGGCTGGGAAAGCA GCATCGGTCT GACCGCCCCC GTAATGTCTG 1151 CCGTTGCCAT TGCCACCGTATCGGTACGCC TTTTCTTTAA AAAAACCCAA 1201 TCTTTACAAA GGAACCCGTC ATGA

This corresponds to the amino acid sequence <SEQ ID 802; ORF125-1>:

1 MSGNASSPSS SSAIGLIWFG AAVSIAEIST GTLLAPLGWQ RGLAALLLGH 51 AVGGALFFAAAYIGALTGRS SMESVRLSFG KRGSVLFSVA NMLQLAGWTA 101 VMIYAGATVS SALGKVLWDGESFVWWALAN GALIVLWLVF GARKTGGLKT 151 VSMLLMLLAV LWLSAEVFST AGSTAAQVSDGMSFGTAVEL SAVMPLSWLP 201 LAADYTRHAR RPFAATLTAT LAYTLTGCWM YALGLAAALFTGETDVAKIL 251 LGAGLGAAGI LAVVLSTVTT TFLDAYSAGA SANNISARFA ETPVAVGVTL301 IGTVLAVMLP VTEYENFLLL IGSVFAPMAA VLIADFFVLK RREEIEGFDF 351AGLVLWLAGF ILYRFLLSSG WESSIGLTAP VMSAVAIATV SVRLFFKKTQ 401 SLQRNPS*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF125 shows 76.5% identity over a 51 as overlap with an ORF (ORF125a)from strain A of N. meningitidis:

The ORF125a partial nucleotide sequence <SEQ ID 803> is:

1 ATGTCGGGCA ATGCCTCCTC TCNTTCATCT TCCGCCGCCA TCGGGCTGAT 51 TTGGTTCGGCGCGGCGGTAT CGATTGCCGA AATCAGCACG GGTACACTGC 101 TTGCGCCTTT GGGCTGGCAGCGCGGTCTGG CNGCTCTGCT TTTGGGTCAT 151 GCCGTCGGCG GCGCGCTGTT TTTTGCGGCGGCGTATATCG GCGCACTGAC 201 CGGACNCANC TCGATGGAAA GCGTGCGCCT GTCGTTCGGCAAACGCGGTT 251 CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG CTGGACGGCG301 GTGATGATTT ACGCCGGCGC AACGGTCAGC TCCGCTTTGG GCAAAGTGTT 351GTGGGACGGC GAATCTTTTG TCTGGTGGGC ATTGGCAAAC GGCGCGCTGA 401 TTGTGCTGTGGCTGGTTTTC GGCGCACGCA AAACAGGCGG GCTGAAAACC 451 GTTTCGATGC TGCTGATGCTGTTGGCGGTT CTGTGGCTGA GTGCCGAANT 501 NTTTTCCACG GCAGGCAGCA CCGCCGCANNGGTNNCAGAC GGCATGAGTT 551 TCGGAACGGC AGTCGAGCTG TCCGCCGTNA TGCCGCTTTCTTGGCTGCCG 601 CTGGCCGCCG ACTACACGCG CCACGCGCGC CGCCCGTTTG CGGCAACCCT651 GACGGCAACG CTCGCCTACA CGCTGACCGG CTGCTGGATG TATGCCTTGG 701GTTTGGCAGC GGCGTTGTTC ACCGGAGAAA CCGACGTGGC AAAAATCCTG 751 CTGGGCGCAGGTTTGGGTGC GGCAGGCATT TTGGCGGTCG TCCTGTCGAC 801 CGTTACCACC ACTTTTCTCGATGCNTACTC CGCCGGCGTA AGTGCCAACA 851 ATATTTCCGC CAAACTTTCG GAAATACCNATCGCCGTTGC CGTCGCCGTT 901 GTCGGCACAC TGCTTGCCGT CCTCCTGCCC GTTACCGAATATGAAAACTT 951 CCTGCTGCTT ATCGGCTCGG TATTTGCGCC GATGGCGGCG GTTTTGATTG1001 CCGACTTTTT CGTCTTGAAA CGGCGTGAGG AGATTGAAGG C..

This encodes a protein having the partial amino acid sequence <SEQ ID804>:

1 MSGNASSXSS SAAIGLIWFG AAVSIAEIST GTLLAPLGWQ RGLAALLLGH 51 AVGGALFFAAAYIGALTGXX SMESVRLSFG KRGSVLFSVA NMLQLAGWTA 101 VMIYAGATVS SALGKVLWDGESFVWWALAN GALIVLWLVF GARKTGGLKT 151 VSMLLMLLAV LWLSAEXFST AGSTAAXVXDGMSFGTAVEL SAVMPLSWLP 201 LAADYTRHAR RPFAATLTAT LAYTLTGCWM YALGLAAALFTGETDVAKIL 251 LGAGLGAAGI LAVVLSTVTT TFLDAYSAGV SANNISAKLS EIPIAVAVAV301 VGTLLAVLLP VTEYENFLLL IGSVFAPMAA VLIADFFVLK RREEIEG..

ORF125a and ORF125-1 show 94.5% identity in 347 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF125 shows 86.2% identity over a 65aa overlap with a predicted ORF(ORF125ng) from N. gonorrhoeae:

An ORF125ng nucleotide sequence <SEQ ID 805> was predicted to encode aprotein having amino acid sequence <SEQ ID 806>:

1 MSGNASSPSS SAAIGLVWFG AAVSIAEIST GTLLAPLGWQ RGLAALLLGH 51 AVGGALFFAAAYIGALTGRS SMESVRLSFG KCGSVLFSVA NMLQLAGWTA 101 VMIYVGATVS SALGKVLWDGESFVWWALAN GALIVLWLVF GARRTGGLKT 151 VSMLLMLLAV LWLSVEVFAS SGTNAAPAVSDGMTFGTAVE LSAVMPLSWL 201 PLAADYTRQA RRPFAATLTA TLAYTLTGCW MYALGLAAALFTGETDVAKI 251 LLGAGLGITG ILAVVLSTVT TTFLDTYSAG ASANNISARF AEIPVAVGVT301 LIRTVLAVML PVTEYKNFLL LIRSVFGPMA GGFDCRLFCL KTA*

Further work revealed the following gonococcal DNA sequence <SEQ ID807>:

1 ATGTCGGGCA ATGCCTCCTC TCCTTCATCT TCCGCCGCCA TCGGGCTGGT 51 TTGGTTCGGCGCGGCGGTAT CGATTGCCGA AATCAGCACG GGTACGCTGC 101 TCGCCCCCTT GGGCTGGCAGCGCGGTCTGG CGGCCCTGCT TTTGGGTCAT 151 GCCGTCGGCG GCGCGCTGTT TTTTGCGGCGGCGTATATCG GCGCACTGAC 201 CGGACGCAGC TCGATGGAAA GTGTGCGCCT GTCGTTCGGCAAATGCGGTT 251 CAGTGCTGTT TTCCGTGGCG AATATGCTGC AACTGGCCGG CTGGACGGCG301 GTGATGATTT ACGTCGGCGC AACGGTCAGC TCCGCTTTGG GCAAAGTGTT 351GTGGGACGGC GAATCCTTTG TCTGGTGGGC ATTGGCAAAC GGCGCACTGA 401 TCGTGCTGTGGCTGGTTTTC GGCGCACGCA GAACGGGCGG GCTGAAAACC 451 GTTTCGATGC TGCTGATGCTGCTTGCCGTG TTGTGGTTGA GCGTCGAAGT 501 GTTCGCTTCG TCCGGCACAA ACGCCGCGCCCGCCGTTTCA GACGGCATGA 551 CCTTCGGAAC GGCAGTCGAA CTGTCCGCCG TCATGCCGCTTTCCTGGCTG 601 CCGCTGGCCG CCGACTACAC GCGCCAAGCA CGCCGCCCGT TTGCGGCAAC651 CCTGACGGCA ACGCTCGCCT ATACGCTGAC GGGCTGCTGG ATGTATGCCT 701TGGGTTTGGC GGCGGCTCTG TTTACCGGAG AAACCGACGT GGCGAAAATC 751 CTGTTGGGCGCGGGCTTGGG CATAACGGGC ATTCTGGCAG TCGTCCTCTC 801 CACCGTTACC ACAACGTTTCTCGATACCTA TTCCGCCGGC GCGAGTGCGA 851 ACAACATTTC CGCGCGTTTT GCGGAAATACCCGTCGCTGT CGGCGTTACC 901 CTGATCGGCA CGGTGCTTGC CGTCATGCTG CCCGTTACCGAATATAAAAA 951 CTTCCTGCTG CTTATCGGCT CGGTATTTGC GCCGATGGCG GCGGTTTTGA1001 TTGCCGACTT TTTCGTCTTA AAACGGCGTG AGGAGATTGA AGGCTTTGAC 1051TTTGCCGGAC TGGTTCTGTG GCTGGCAGGC TTCATCCTCT ACCGCTTCCT 1101 GCTCTCGTCCGGTTGGGAAA GCAGCATCGG TCTGACCGCC CCCGTAATGT 1151 CTGCCGTTGC CATTGCCACCGTATCGGTAC GCCTTTTCTT TAAAAAAACC 1201 CAATCTTTAC AAAGGAACCC GTCATGA

This corresponds to the amino acid sequence <SEQ ID 808; ORF125ng-1>:

1 MSGNASSPSS SAAIGLVWFG AAVSIAEIST GTLLAPLGWQ RGLAALLLGH 51 AVGGALFFAAAYIGALTGRS SMESVRLSFG KCGSVLFSVA NMLQLAGWTA 101 VMIYVGATVS SALGKVLWDGESFVWWALAN GALIVLWLVF GARRTGGLKT 151 VSMLLMLLAV LWLSVEVFAS SGTNAAPAVSDGMTFGTAVE LSAVMPLSWL 201 PLAADYTRQA RRPFAATLTA TLAYTLTGCW MYALGLAAALFTGETDVAKI 251 LLGAGLGITG ILAVVLSTVT TTFLDTYSAG ASANNISARF AEIPVAVGVT301 LIGTVLAVML PVTEYKNFLL LIGSVFAPMA AVLIADFFVL KRREEIEGFD 351FAGLVLWLAG FILYRFLLSS GWESSIGLTA PVMSAVAIAT VSVRLFFKKT 401 QSLQRNPS*

ORF125ng-1 and ORF125-1 show 95.1% identity in 408 aa overlap:

Based on this analysis, including the presence of putative leadersequence and transmembrane domains in the gonococcal protein, it ispredicted that the proteins from N. meningitidis and N. gonorrhoeae, andtheir epitopes, could be useful antigens for vaccines or diagnostics, orfor raising antibodies.

Example 96

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 809>:

1 ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCGGGAA GGCTGACCGC 51 GTTGCAGCTTGCAGAACAAG GTTATCAGAT TGCACTTTTC GATAAAAGCT 101 GCCGCCGGGG CGAACACGCCGCCGCCTATG TAGCCGCCGC CATGCTCGCG 151 CCTGCAGCGG A.ACGGTCGA AGCCACGCCCGAAGTGGTCA GGCTGGGCAG 201 GCAGAGCATC CCGCTTTGGC GCGGCATCCG ATGCCGTCTGAACACGCACA 251 CGATGATGCA GGAAAACGGC AGCCTGATTG TATGGCACGG GCAGGACAAG301 CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG GCGT.ACGGA 351TGACGAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA CGCGAACCGC 401 AACTCGGCGGACGTTTTTAA GACGGCATCT ACCTGCCGAC CGAAGC.CAG 451 CTCGACGGGC GGCAATTATAGTCTGCACTT GCCGACGCTT TGGACGAACT 501 GAACGTCCCC TGCCATTGGG AACACGAATGCGTCCCCGAA GCCTGCAAG..

This corresponds to the amino acid sequence <SEQ ID 810; ORF126>:

1 MTRIAILGGG LSGRLTALQL AEQGYQIALF DKSCRRGEHA AAYVAAAMLA 51 PAAXTVEATPEVVRLGRQSI PLWRGIRCRL NTHTMMQENG SLIVWHGQDK 101 PLSSEFVRHL KRGGXTDDEIVRWRADDIAE REPQLGGRFX DGIYLPTEXQ 151 LDGRQLXSAL ADALDELNVP CHWEHECVPEACK...

Further work revealed the complete nucleotide sequence <SEQ ID 811>:

1 ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCGGGAA GGCTGACCGC 51 GTTGCAGCTTGCAGAACAAG GTTATCAGAT TGCACTTTTC GATAAAGGCT 101 GCCGCCGGGG CGAACACGCCGCCGCCTATG TTGCCGCCGC CATGCTCGCG 151 CCTGCGGCGG AAGCGGTCGA AGCCACGCCCGAAGTGGTCA GGCTGGGCAG 201 GCAGAGCATC CCGCTTTGGC GCGGCATCCG ATGCCGTCTGAACACGCACA 251 CGATGATGCA GGAAAACGGC AGCCTGATTG TGTGGCACGG GCAGGACAAG301 CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG GCGTAGCGGA 351TGACGAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA CGCGAACCGC 401 AACTCGGCGGACGTTTTTCA GACGGCATCT ACCTGCCGAC CGAAGGCCAG 451 CTCGACGGGC GGCAAATATTGTCTGCACTT GCCGACGCTT TGGACGAACT 501 GAACGTCCCC TGCCATTGGG AACACGAATGCGTCCCCGAA GGCCTGCAAG 551 CCCAATACGA CTGGCTGATC GACTGCCGCG GCTACGGCGCAAAAACCGCG 601 TGGAACCAAT CCCCCGAGCA CACCAGCACC CTGCGCGGCA TACGCGGCGA651 AGTGGCGCGG GTTTACACAC CCGAAATCAC GCTCAACCGC CCCGTGCGTC 701TGCTCCATCC GCGTTATCCG CTCTACATCG CCCCGAAAGA AAACCACGTC 751 TTCGTCATCGGCGCGACCCA AATCGAAAGC GAAAGCCAAG CCCCCGCCAG 801 CGTGCGTTCA GGGTTGGAACTCTTGTCCGC ACTCTATGCC ATCCACCCCG 851 CCTTCGGCGA AGCCGACATC CTCGAAATCGCCACCGGCCT GCGCCCCACG 901 CTCAACCACC ACAACCCCGA AATCCGTTAC AACCGCGCCCGACGCCTGAT 951 TGAAATCAAC GGCCTTTTCC GCCACGGTTT CATGATCTCC CCCGCCGTAA1001 CCGCCGCCGC CGCCAGATTG GCAGTGGCAC TGTTTGACGG AAAAGACGCG 1051CCCGAACGCG ATAAAGAAAG CGGTTTGGCG TATATCCGAA GACAAGATTA 1101 A

This corresponds to the amino acid sequence <SEQ ID 812; ORF126-1>:

1 MTRIAILGGG LSGRLTALQL AEQGYQIALF DKGCRRGEHA AAYVAAAMLA 51 PAAEAVEATPEVVRLGRQSI PLWRGIRCRL NTHTMMQENG SLIVWHGQDK 101 PLSSEFVRHL KRGGVADDEIVRWRADDIAE REPQLGGRFS DGIYLPTEGQ 151 LDGRQILSAL ADALDELNVP CHWEHECVPEGLQAQYDWLI DCRGYGAKTA 201 WNQSPEHTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYPLYIAPKENHV 251 FVIGATQIES ESQAPASVRS GLELLSALYA IHPAFGEADI LEIATGLRPT301 LNHHNPEIRY NRARRLIEIN GLFRHGFMIS PAVTAAAARL AVALFDGKDA 351PERDKESGLA YIRRQD*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF126 shows 90.0% identity over a 180aa overlap with an ORF (ORF126a)from strain A of N. meningitidis:

The complete length ORF126a nucleotide sequence <SEQ ID 813> is:

1 ATGACCCGTA TCGCCATCCT CGGCGGCGGC CTCTCNGGAA GGCTGACCGC 51 ACTGCAGCTTGCAGAACAAG GTTATCAGAT TGCACTTTTC GATAAAGGCT 101 GCCGCCGGGG CGAACACGCCGCCGCCTATG TTGCCGCCGC CATGCTCGCG 151 CCTGCGGCGG AAGCGGTCGA AGCCACGCCTGAAGTGGTCA GGCTGGGCAG 201 GCAGANCATC CCGCTTTGGC GCGGCATCCG ATGCCATCTGAAAACGCCTG 251 CCATGATGCA NGAAAACGGC AGCCTGATTG TGTGGCACGG GCAGGACAAA301 CCTTTATCCA ACGAGTTCGT CCGCCATCTC AAACGCGGCG GCGTAGCGGA 351TGACNAAATC GTCCGTTGGC GCGCCGACGA CATCGCCGAA CGCGAACCGC 401 AACTCGGCGGACGTTTTTCA GACGGCATCT ACCTGCCGAC CGAAGGCCAG 451 CTCGACGGGC GGCAAATATTGTCTGCACTT GCCGACGCTT TGGACGAACT 501 GAACGTCCCC TGCCATTGGG AACACGAATGTGCCCCCGAA GACTTGCAAG 551 CCCAATACGA CTGGCTGATC GACTGCCGCG GCTACGGCGCAAAAACCGCG 601 TGGAACCAAT CCCCCGANNA NACCAGCACC CTGCGCGGCA TACGCGGCGA651 AGTGGCGCGG GTTTACACAC CCGAAATCAC GCTCAACCGC CCCGTGCGCC 701TGCTACACCC GCGCTATCCG CTNTACATCG CCCCGAAAGA AAACCNCGTC 751 TTCGTCATCGGCGCGACCCA AATCGAAAGC GAAAGCCAAG CACCTGCCAG 801 CGTGCGTTCC GGGCTGGAACTCTTATCCGC ACTCTATGCC GTCCACCCCG 851 CCTTCGGCGA AGCCGACATC CTCGAAATCGCCACCGGCCT GCGCCCCACG 901 CTCAATCACC ACAACCCCGA AATCCGTTAC AACCGCGCCCGACGCCTGAT 951 TGAAATCAAC GGCCTTTTCC GCCACGGTTT CATGATCTCC CCCGCCGTAA1001 CCGCCGCCGC CGTCAGATTG GCAGTGGCAC TGTTTGACGG AAAAGANGCG 1051CCCGAACGCG ATGAAGAAAG CGGTTTGGCG TATATCCGAA GACAAGATTA 1101 A

This encodes a protein having amino acid sequence <SEQ ID 814>:

1 MTRIAILGGG LSGRLTALQL AEQGYQIALF DKGCRRGEHA AAYVAAAMLA 51 PAAEAVEATPEVVRLGRQXI PLWRGIRCHL KTPAMMXENG SLIVWHGQDK 101 PLSNEFVRHL KRGGVADDXIVRWRADDIAE REPQLGGRFS DGIYLPTEGQ 151 LDGRQILSAL ADALDELNVP CHWEHECAPEDLQAQYDWLI DCRGYGAKTA 201 WNQSPXXTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYPLYIAPKENXV 251 FVIGATQIES ESQAPASVRS GLELLSALYA VHPAFGEADI LEIATGLRPT301 LNHHNPEIRY NRARRLIEIN GLFRHGFMIS PAVTAAAVRL AVALFDGKXA 351PERDEESGLA YIRRQD*

ORF126a and ORF126-1 show 95.4% identity in 366 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF126 shows 90% identity over a 180 aa overlap with a predicted ORF(ORF126ng) from N. gonorrhoeae:

An ORF126ng nucleotide sequence <SEQ ID 815> was predicted to encode aprotein having amino acid sequence <SEQ ID 816>:

1 MTRIAVLGGG LSGRLTALQL AEQGYQIELF DKGTRQGEHA AAYVAAAMLA 51 PAAEAVEATPEVIRLGRQSI PLWRGIRCRL NTLTMMQENG SLIVWHGQDK 101 PLSSEFVRHL KRGGVADDEIVRWRADEIAE REPQLGGRFS DGIYLPTEGQ 151 LDGRQILSAL ADALDELNVP CHWEHECAPQDLQAQYDWVI DCRGYGAKTA 201 WNQSPEHTST LRGIRGEVRG FTRPKSRSTA PCACCTRAIRSTSPRKKTTS 251 SSSARPKSKA KAKPPPAYVP GWNSYPRSMP STPPSAKPTS SKWRPGLRPT301 LNHHNPEIRY SRERRLIEIN GLFRHGFMIS PAVTAAAVRL AVALFDGKDA 351PERDEESGLA YIGRQD*

Further work revealed the following gonococcal DNA sequence <SEQ ID817>:

1 ATGACCCGTA TCGCCGTCCT CGGAGGCGGC CTTTCCGGAA GGCTGACCGC 51 ATTGCAGCTTGCAGAACAAG GTTATCAGAT TGAACTTTTC GACAAGGGCA 101 CCCGCCAAGG CGAACACGCCGCCGCCTATG TTGCCGCCGC GATGCTCGCG 151 CCTGCGGCGG AAGCGGTCGA GGCAACGCCCGAAGTCATCA GGCTGGGCAG 201 GCAGAGCATT CCGCTTTGGC GCGGCATCCG ATGCCGTCTGAACACGCTCA 251 CGATGATGCA GGAAAACGGC AGCCTGATTG TGTGGCACGG GCAGGACAAG301 CCATTATCCA GCGAGTTCGT CCGCCATCTC AAACGCGGCG GCGTAGCGGA 351TGACGAAATC GTCCGTTGGC GCGCCGATGA AATCGCCGAA CGCGAACCGC 401 AACTCGGCGGACGTTTTTCA GACGGCATCT ACCTGCCGAC CGAAGGCCAG 451 CTCGACGGGC GGCAAATATTGTCTGCACTT GCCGACGCTT TGGACGAACT 501 GAACGTCCCT TGCCATTGGG AACACGAATGCGCCCCCCAA GACCTGCAAG 551 CCCAATACGA CTGGGTAATC GACTGCCGGG GCTACGGCGCGAAAACCGCG 601 TGGAACCAAT CCCCCGAGCA CACCAGCACC TTGCGCGGCA TACGCGGCGA651 AGTGGCGCGG GTTTACACGC CCGAAATCAC GCTCAACCGC CCCGTGCGCC 701TGCTGCACCC GCGCTATCCG CTCTACATCG CCCCGAAAGA AAACCACGTC 751 TTCGTCATCGGCGCGACCCA AATCGAAAGC GAAAGCCAAG CCCCCGCCAG 801 CGTACGTTCC GGGCTGGAACTCTTATCCGC GCTCTATGCC GTCCACCCCG 851 CCTTCGGCGA AGCCGACATC CTCGAAATCGCCGCCGGCCT GCGCCCCACG 901 CTCAACCACC ACAACCCCGA AATCCGCTAC AGCCGCGAACGCCGCCTCAT 951 CGAAATCAAC GGCCTTTTCC GGCACGGCTT TATGATTTCC CCCGCCGTAA1001 CCGCCGCCGC CGTCAGATTG GCAGTGGCAC TGTTTGACGG AAAAGACGCG 1051CCCGAACGTG ATGAAGAAAG CGGTTTGGCG TATATCGGAA GACAAGATTA 1101 A

This corresponds to the amino acid sequence <SEQ ID 818; ORF126ng-1>:

1 MTRIAVLGGG LSGRLTALQL AEQGYQIELF DKGTRQGEHA AAYVAAAMLA 51 PAAEAVEATPEVIRLGRQSI PLWRGIRCRL NTLTMMQENG SLIVWHGQDK 101 PLSSEFVRHL KRGGVADDEIVRWRADEIAE REPQLGGRFS DGIYLPTEGQ 151 LDGRQILSAL ADALDELNVP CHWEHECAPQDLQAQYDWVI DCRGYGAKTA 201 WNQSPEHTST LRGIRGEVAR VYTPEITLNR PVRLLHPRYPLYIAPKENHV 251 FVIGATQIES ESQAPASVRS GLELLSALYA VHPAFGEADI LEIAAGLRPT301 LNHHNPEIRY SRERRLIEIN GLFRHGFMIS PAVTAAAVRL AVALFDGKDA 351PERDEESGLA YIGRQD*

ORF126ng-1 and ORF126-1 show 95.1% identity in 366 aa overlap:

Furthermore, ORF126ng-1 shows homology to a putative Rhizobium oxidaseflavoprotein:

gi|2627327 (AF004408) putative amino acid oxidase flavoprotein[Rhizobium etli] Length = 327 Score = 169 bits (423), Expect = 3e−41Identities = 112/329 (34%), Positives = 163/329 (49%), Gaps = 25/329(7%) Query: 3RIAVLGGGLSGRLTALQLAEQGYQIELFDKGTRQGEHXXXXXXXXXXXXXXXXXXXXXXX 62 RI V GG++G   A QL   G+++ L ++    G Sbjct: 2RILVNGAGVAGLTVAWQLYRHGFRVTLAERAGTVGA-GASGFAGGMLAPWCERESAEEPV 60 Query:63 IRLGRQSIPLWRGIRCRLNTLTMMQENGSLIVWHGQDKPLSSEFVRHLKRGGVADDEIVR 122+ LGR +   W            +   G+L+V  G+D      F R    G    DE+ Sbjct: 61LTLGRLAADWWEAA-----LPGHVHRRGTLVVAGGRDTGELDRFSRRTS-GWEWLDEVA- 113 Query:123 WRADEIAEREPQLGGRFSDGIYLPTEGQLDGRQILSALADALDELNVPCHWEHECAPQDL 182     IA  EP L GRF   ++   E  LD RQ L+ALA  L++  +           + Sbjct: 114-----IAALEPDLAGRFRRALFFRQEAHLDPRQALAALAAGLEDARMRLTLG---VVGES 165 Query:183 QAQYDWVIDCRGYGAKTAWNQSPEHTSTLRGIRGEVARVYTPEITLNRPVRLLHPRYPLY 242   +D V+DC G                LRG+RGE+  V T E++L+RPVRLLHPR+P+Y Sbjct: 166DVDHDRVVDCTGAA-------QIGRLPGLRGVRGEMLCVETTEVSLSRPVRLLHPRHPIY 218 Query:243 IAPKENHVFVIGATQIESESQAPASVRSGLELLSALYAVHPAFGEADILEIAAGLRPTLN 302 IP++ + F++GAT IES+   P + RS +ELL+A YA+HPAFGEA + E  AG+RP Sbjct: 219IVPRDKNRFMVGATMIESDDGGPITARSLMELLNAAYAMHPAFGEARVTETGAGVRPAYP 278 Query:303 HHNPEIRYSRERRLIEINGLFRHGFMISP 331  + P  R ++E R + +NGL+RHGF+++PSbjct: 279 DNLP--RVTQEGRTLHVNGLYRHGFLLAP 305

This analysis suggests that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 97

The following DNA sequence, believed to be complete, was identified inN. meningitidis <SEQ ID 819>:

1 ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT CAGTGGTCTT 51 GATATTGTCTGTACTTGCTT TAATTGTTTA TCCGAGCTAT CGCAATTATG 101 TTGAGAAAGC AAAGATAAATGCAGTGCGGG CAGCCTTGTT AGAAAATGCA 151 CATTTTATGG AAAAGTTTTA TCTGCAGAATGGGAGGTTTA AACAAACATC 201 TACCAAGTGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGCTTTTGTATCC 251 GTTTGAATGG AATCGtCGCG CGGG..GCTT TAGACAGTAA ATTCATGTTG301 AAGGCGGTAG CCATAGATAA AGATAAAAAT CCTTTTATTA TTAAGATGAA 351TGAAAATCTA GTAACCTTTA aTTTGCAAGA AGTCCGCCAG TTCGTGTAGT 401 GACGGGCTGGATTATTTTAA AGGAAATGAT AAGGACTGCA AGTTACTTAA 451 GTAG

This corresponds to the amino acid sequence <SEQ ID 820; ORF127>:

1 MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN AVRAALLENA 51 HFMEKFYLQNGRFKQTSTKW PSLPIKEAEG FCIRLNGIVA RXALDSKFML 101 KAVAIDKDKN PFIIKMNENLVTFICKKSAS SCSDGLDYFK GNDKDCKLLK 151 *

Further work revealed the following DNA sequence <SEQ ID 821>:

1 ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT CAGTGGTCTT 51 GATATTGTCTGTACTTGCTT TAATTGTTTA TCCGAGCTAT CGCAATTATG 101 TTGAGAAAGC AAAGATAAATGCAGTGCGGG CAGCCTTGTT AGAAAATGCA 151 CATTTTATGG AAAAGTTTTA TCTGCAGAATGGGAGGTTTA AACAAACATC 201 TACCAAGTGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGCTTTTGTATCC 251 GTTTGAATGG AATCGCGCGC GGGGCTTTAG ACAGTAAATT CATGTTGAAG301 GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA AGATGAATGA 351AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG TGTAGTGACG 401 GGCTGGATTATTTTAAAGGA AATGATAAGG ACTGCAAGTT ACTTAAGTAG

This corresponds to the amino acid sequence <SEQ ID 822; ORF127-1>:

1 MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN AVRAALLENA 51 HFMEKFYLQNGRFKQTSTKW PSLPIKEAEG FCIRLNGIAR GALDSKFMLK 101 AVAIDKDKNP FIIKMNENLVTFICKKSASS CSDGLDYFKG NDKDCKLLK*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF127 shows 98.0% identity over a 150aa overlap with an ORF (ORF127a)from strain A of N. meningitidis:

The complete length ORF127a nucleotide sequence <SEQ ID 823> is:

1 ATGACTGATA ATCGGGGGTT TACGCTGGTT GAATTAATAT CAGTGGTCTT 51 GATATTGTCTGTACTTGCTT TAATTGTTTA TCCGAGCTAT CGCAATTATG 101 TTGAGAAAGC AAAGATAAATACAGTGCGGG CAGCCTTGTT AGAAAATGCA 151 CATTTTATGG AAAAGTTTTA TCTGCAGAATGGGAGATTTA AACAAACATC 201 TACCAAATGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGCTTTTGTATCC 251 GTTTGAATGG AATCGCGCGC GGGGCCTTAG ACAGTAAATT CATGTTGAAG301 GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA AGATGAATGA 351AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG TGTAGTGACG 401 GGCTGGATTATTTTAAAGGA AATGATAAGG ACTGCAAGTT ACTTAAGTAG

This encodes a protein having amino acid sequence <SEQ ID 824>:

1 MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN TVRAALLENA 51 HFMEKFYLQNGRFKQTSTKW PSLPIKEAEG FCIRLNGIAR GALDSKFMLK 101 AVAIDKDKNP FIIKMNENLVTFICKKSASS CSDGLDYFKG NDKDCKLLK*

ORF127a and ORF127-1 show 99.3% identity in 149 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF127 shows 97.3% identity over a 150 aa overlap with a predicted ORF(ORF127ng) from N. gonorrhoeae:

The complete length ORF127ng nucleotide sequence <SEQ ID 825> is:

1 ATGACTGATA ATCGGGGGTT TACACTGGTT GAATTAATAT CAGTGGTCTT 51 GATATTGTCTGTACTTGCTT TAATTGTTTA TCCGAGCTAT CGCAATTATG 101 TTGAGAAAGC AAAGATAAATGCAGTGCGGG CAGCCTTGTT AGAAAATGCA 151 CATTTTATGG AAAAGTTTTA TCTGCAGAATGGGAGATTTA AACAAACATC 201 TACCAAATGG CCAAGTTTGC CGATTAAAGA GGCAGAAGGCTTTTGTATCC 251 GTTTGAATGG AATCGCGCGC GGGGCTTTAG ACAGTAAATT CATGTTGAAG301 GCGGTAGCCA TAGATAAAGA TAAAAATCCT TTTATTATTA AGATGAATGA 351AAATCTAGTA ACCTTTATTT GCAAGAAGTC CGCCAGTTCG TGTAGTGACG 401 GGCTGGATTATTTTAAAGGA AATGATAAGG ACTGCAAGTT ACTTAAGTAG

This encodes a protein having amino acid sequence <SEQ ID 826>:

1 MTDNRGFTLV ELISVVLILS VLALIVYPSY RNYVEKAKIN AVRAAFLENA 51 HFMEKFYLQNGRFKQTSTKW PSLPIKEAEG FCIRLNGIAR GALDSKFMLK 101 AVAIDKDKNP FIIKMNENLVTFICKKSASS CSDRLDYFKG NDKDCKLLK*

ORF127ng and ORF127-1 show 100.0% identity in 149 aa overlap:

This analysis, including the fact that the predicted transmembranedomain is shared by the meningococcal and gonococcal proteins, suggeststhat the proteins from N. meningitidis and N. gonorrhoeae, and theirepitopes, could be useful antigens for vaccines or diagnostics, or forraising antibodies.

Example 98

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 827>

1 ..GTGTCGCTGG CTTCGGTGAT TGCCTCTCAA ATCTTCCTTT ACGAAGATTT 51  CAACCAAATG CGGAAAACCC GTGGAGCTAT CTGCGGTTTT CTTGTCCAAT 101  ATTTATCTGG GGTTTCAGCA GGGGTATTTC GATTTGAGTG CCGACGAGAA 151  CCCCGTACTG CATATCTGGT CTTTGGCAGT AGAGGAACAG TATTACCTCC 201  TGTATCCCCT TTTGCTGATA TTTTGCTGCA AAAAAACCAA ATCGCTACGG 251  GTGCTGCGTA ACATCAGCAT CATCCTGTTT TTGATTTTGA CTGCCTCATC 301  GTTTTTGCCA AGCGGGTTTT ATACCGACAT CCTCAACCAA CCCAATACTT 351  ATTACCTTTC GACACTGAGG TTTCCCGAGC TGTTGGCAGG TTCGCTGCTG 401  GCGGTTTACG GGCAAACGCA AAACGGCAGA CGGCAAACAG CAAATGGAAA 451  ACGGCAGTTG CTTTCATCAC TCTGCTTCGG CGCATTGCTT GCCTGCCTGT 501  TCGTGATTGA CAAACACAAT CCGTTTATCC CGGGAATGAC CCTGCTCCTT 551  CCCTGCCTGC TGACGGCACT GCTTATCCGG AGTATGCAAT ACGGGACACT 601  TCCGACCCGC ATCCTGTCGG CAAGCCCCAT CGTATTTGTC GGCAAAATCT 651  CTTATTCCCT ATACCTGTAC CATTGGATTT TTATTGCTTT CGCTCCGCTC 701  ATTAGAGGCG GGAAACAGCT CGGACTGCCT GCCG..

This corresponds to the amino acid sequence <SEQ ID 828; ORF128>:

1 ..VSLASVIASQ IFLYEDFNQM RKTVELSAVF LSNIYLGFQQ GYFDLSADEN 51  PVLHIWSLAV EEQYYLLYPL LLIFCCKKTK SLRVLRNISI ILFLILTASS 101  FLPSGFYTDI LNQPNTYYLS TLRFPELLAG SLLAVYGQTQ NGRRQTANGK 151  RQLLSSLCFG ALLACLFVID KHNPFIPGMT LLLPCLLTAL LIRSMQYGTL 201  PTRILSASPI VFVGKISYSL YLYHWIFIAF APLIRGGKQL GLPA..

Further work revealed the complete nucleotide sequence <SEQ ID 829>:

1 ATGCAAGCTG TCCGATACAG ACCGGAAATT GACGGATTGC GGGCCGTCGC 51 CGTGCTATCCGTCATGATTT TCCACCTGAA TAACCGCTGG CTGCCCGGAG 101 GATTCCTGGG GGTGGACATTTTCTTTGTCA TCTCAGGATT CCTCATTACC 151 GGCATCATTC TTTCTGAAAT ACAGAACGGTTCTTTTTCTT TCCGGGATTT 201 TTATACCCGC AGGATTAAGC GGATTTATCC TGCCTTTATTGCGGCCGTGT 251 CGCTGGCTTC GGTGATTGCC TCTCAAATCT TCCTTTACGA AGATTTCAAC301 CAAATGCGGA AAACCGTGGA GCTTTCTGCG GTTTTCTTGT CCAATATTTA 351TCTGGGGTTT CAGCAGGGGT ATTTCGATTT GAGTGCCGAC GAGAACCCCG 401 TACTGCATATCTGGTCTTTG GCAGTAGAGG AACAGTATTA CCTCCTGTAT 451 CCCCTTTTGC TGATATTTTGCTGCAAAAAA ACCAAATCGC TACGGGTGCT 501 GCGTAACATC AGCATCATCC TGTTTTTGATTTTGACTGCC TCATCGTTTT 551 TGCCAAGCGG GTTTTATACC GACATCCTCA ACCAACCCAATACTTATTAC 601 CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GCAGGTTCGC TGCTGGCGGT651 TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGCAAAT GGAAAACGGC 701AGTTGCTTTC ATCACTCTGC TTCGGCGCAT TGCTTGCCTG CCTGTTCGTG 751 ATTGACAAACACAATCCGTT TATCCCGGGA ATGACCCTGC TCCTTCCCTG 801 CCTGCTGACG GCACTGCTTATCCGGAGTAT GCAATACGGG ACACTTCCGA 851 CCCGCATCCT GTCGGCAAGC CCCATCGTATTTGTCGGCAA AATCTCTTAT 901 TCCCTATACC TGTACCATTG GATTTTTATT GCTTTCGCCCATTACATTAC 951 AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT GCCGCGTTGA1001 CGGCCGGATT TTCCCTGTTG AGTTATTATT TGATTGAACA GCCGCTTAGA 1051AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTCT ATCTCGCCCC 1101 GTCCCTGATACTTGTCGGTT ACAACCTGTA CGCAAGGGGG ATATTGAAAC 1151 AGGAACACCT CCGCCCGTTGCCCGGCGCGC CCCTTGCTGC GGAAAATCAT 1201 TTTCCGGAAA CCGTCCTGAC CCTCGGCGACTCGCACGCCG GACACCTGAG 1251 GGGGTTTCTG GATTATGTCG GCAGCCGGGA AGGGTGGAAAGCCAAAATCC 1301 TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TAGATGAGAA GCTGGCAGAC1351 AACCCGTTAT GTCGAAAATA CCGGGATGAA GTTGAAAAAG CCGAAGCCGT 1401TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG CCTGTGCCGA 1451 GATTTGAAGCGCAATCCTTC CTAATACCCG GGTTCCCAGC CCGATTCAGG 1501 GAAACCGTCA AAAGGATAGCCGCCGTCAAA CCCGTCTATG TTTTTGCAAA 1551 CAACACATCA ATCAGCCGTT CGCCCCTGAGGGAGGAAAAA TTGAAAAGAT 1601 TTGCCGCAAA CCAATATCTC CGCCCCATTC AGGCTATGGGCGACATCGGC 1651 AAGAGCAATC AGGCGGTCTT TGATTTGATT AAAGATATTC CCAATGTGCA1701 TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC GAAATATACG 1751GCCGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT CGGTTCTTAT 1801 TATATGGGGCGGGAATTCCA CAAACACGAA CGCCTGCTTA AATCTTCCCA 1851 CGGCGGCGCA TTGCAGTAG

This corresponds to the amino acid sequence <SEQ ID 830; ORF128-1>:

1 MQAVRYRPEI DGLRAVAVLS VMIFHLNNRW LPGGFLGVDI FFVISGFLIT 51 GIILSEIQNGSFSFRDFYTR RIKRIYPAFI AAVSLASVIA SQIFLYEDFN 101 QMRKTVELSA VFLSNIYLGFQQGYFDLSAD ENPVLHIWSL AVEEQYYLLY 151 PLLLIFCCKKTKSLRVLRNI SIILFLILTA SSFLPSGFYT DILNQPNTYY 201 LSTLRFPELL AGSLLAVYGQTQNGRRQTAN GKRQLLSSLC FGALLACLFV 251 IDKHNPFIPG MTLLLPCLLT ALLIRSMQYGTLPTRILSAS PIVFVGKISY 301 SLYLYHWIFI AFAHYITGDK QLG LPAVSAV AALTAGFSLLSYYLIEQPLR 351 KRKMTFKKAF FCLYLAPSLI LVGYNLYARG ILKQEHLRPL PGAPLAAENH401 FPETVLTLGD SHAGHLRGFL DYVGSREGWK AKILSLDSEC LVWVDEKLAD 451NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF LIPGFPARFR 501 ETVKRIAAVKPVYVFANNTS ISRSPLREEK LKRFAANQYL RPIQAMGDIG 551 KSNQAVFDLI KDIPNVHWVDAQKYLPKNTV EIYGRYLYGD QDHLTYFGSY 601 YMGREFHKHE RLLKSSHGGA LQ*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with Hypothetical Integral Membrane Protein H10392 of H.influenzae (Accession Number U32723)

ORF128 and HI0392 show 52% aa identity in 180aa overlap:

Orf128: 1 VSLASVIASQIFLYEDFNQMRKTVELSAVFLSNIYLGFQQGYFDLSADENPVLHIWSLAV60 ++L S IAS IF+Y DFN++RKT+EL+  FLSN YLG  QGYFDLSA+ENPVLHIWSLAV HI0392:46 MALVSFIASAIFIYNDFNKLRKTIELAIAFLSNFYLGLTQGYFDLSANENPVLHIWSLAV 105Orf128: 61 EEQXXXXXXXXXIFCCKKTKSLRVLRNISIILFLILTASSFLPSGFYTDILNQPNTYYLS120 E Q         I   KK + ++VL  I++ILF IL A+SF+ + FY ++L+QPN YYLS HI0392:106 EGQYYLIFPLILILAYKKFREVKVLFIITLILFFILLATSFVSANFYKEVLHQPNIYYLS 165Orf128: 121 TLRFPELLAGSLLAVYGQTQNGRRQTANGKRQLLSSLCFGALLACLFVIDKHNPFIPGMT180  LRFPELL GSLLA+Y    N + Q +     +L+ L    L +CLF+++ +  FIPG+T HI0392:166 NLRFPELLVGSLLAIYHNLSN-KVQLSKQVNNILAILSTLLLFSCLFLMNNNIAFIPGIT 224Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF128 shows 98.0% identity over a 244aa overlap with an ORF (ORF128a)from strain A of N. meningitidis:

The complete length ORF128a nucleotide sequence <SEQ ID 831> is:

1 ATGCAAGCTG TCCGATACAG ACCGGAAATT GACGGATTGC GGGCCGTCGC 51 CGTGCTATCCGTCATGATTT TCCACCTGAA TAACCGCTGG CTGCCCGGAG 101 GATTCCTGGG GGTGGACATTTTCTTTGTCA TCTCAGGATT CCTCATTACC 151 GGCATCATTC TTTCTGAAAT ACAGAACGGTTCTTTTTCTT TCCGGGATTT 201 TTATACCCGC AGGATTAAGC GGATTTATCC TGCTTTTATTGCGGCCGTGT 251 CGCTGGCTTC GGTGATTGCC TCTCAAATCT TCCTTTACGA AGATTTCAAC301 CAAATGCGGA AAACCGTGGA GCTTTCTGCG GTTTTCTTGT CCAATATTTA 351TCTGGGGTTT CAGCAGGGGT ATTTCGATTT GAGTGCCGAC GAGAACCCCG 401 TACTGCATATCTGGTCTTTG GCAGTAGAGG AACAGTATTA CCTCCTGTAT 451 CCTCTTTTGC TGATATTTTGCTGCAAAAAA ACAAAATCGC TACGGGTGCT 501 GCGTAACATC AGCATCATCC TATTTCTGATTTTGACTGCC ACATCGTTTT 551 TGCCAAGCGG GTTTTATACC GATATTCTCA ACCAACCCAATACTTATTAC 601 CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GCAGGTTCGC TGCTGGCGGT651 TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGCAAAT GGAAAACGGC 701AGTTGCTTTC ATCACTCTGC TTCGGCGCAT TGCTTGCCTG CCTGTTCGTG 751 ATTGACAAACACAATCCGTT TATCCCGGGA ATGACCCTGC TCCTTCCCTG 801 CCTGCTGACG GCACTGCTTATCCGGAGTAT GCAATACGGG ACACTTCCGA 851 CCCGCATCCT GTCGGCAAGC CCCATCGTATTTGTCGGCAA AATCTCTTAT 901 TCCCTATACC TGTACCATTG GATTTTTATT GCTTTCGCCCATTACATTAC 951 AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT GCCGCGTTGA1001 CGGCCGGATT TTCCCTGTTG AGTTATTATT TGATTGAACA GCCGCTTAGA 1051AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTCT ATCTCGCCCC 1101 GTCCCTGATACTTGTCGGTT ACAACCTGTA CGCAAGGGGG ATATTGAAAC 1151 AGGAACACCT CCGCCCGTTGCCCGGCGCGC CCCTTGCTGC GGAAAATCAT 1201 TTTCCGGAAA CCGTCCTGAC CCTCGGCGACTCGCACGCCG GACACCTGCG 1251 GGGGTTTCTG GATTATGTCG GCAGCCGGGA AGGGTGGAAAGCCAAAATCC 1301 TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TAGATGAGAA GCTGGCAGAC1351 AACCCGTTAT GTCGAAAATA CCGGGATGAA GTTGAAAAAG CCGAAGCCGT 1401TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG CCCGTGCCGA 1451 GATTTGAAGCGCAATCCTTC CTAATACCCG GGTTCCCAGC CCGATTCAGG 1501 GAAACCGTCA AAAGGATAGCCGCCGTCAAA CCCGTCTATG TTTTTGCAAA 1551 CAACACATCA ATCAGCCGTT CGCCCCTGAGGGAGGAAAAA TTGAAAAGAT 1601 TTGCCGCAAA CCAATATCTC CGCCCCATTC AGGCTATGGGCGACATCGGC 1651 AAGAGCAATC AGGCGGTCTT TGATTTGATT AAAGATATTC CCAATGTGCA1701 TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC GAAATATACG 1751GCCGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT CGGTTCTTAT 1801 TATATGGGGCGGGAATTTCA CAAACACGAA CGCCTGCTTA AATCTTCTCG 1851 CGACGGCGCA TTGCAGTAG

This encodes a protein having amino acid sequence <SEQ ID 832>:

1 MQAVRYRPEI DGLRAVAVLS VMIFHLNNRW LPGGFLGVDI FFVISGFLIT 51 GIILSEIQNGSFSFRDFYTR RIKRIYPAFI AAVSLASVIA SQIFLYEDFN 101 QMRKTVELSA VFLSNIYLGFQQGYFDLSAD ENPVLHIWSL AVEEQYYLLY 151 PLLLIFCCKKTKSLRVLRNI SIILFLILTA TSFLPSGFYT DILNQPNTYY 201 LSTLRFPELL AGSLLAVYGQTQNGRRQTAN GKRQLLSSLC FGALLACLFV 251 IDKHNPFIPG MTLLLPCLLT ALLIRSMQYGTLPTRILSAS PIVFVGKISY 301 SLYLYHWIFI AFAHYITGDK QLGLPAVSAV AALTAGFSLLSYYLIEQPLR 351 KRKMTFKKAF FCLYLAPSLI LVGYNLYARG ILKQEHLRPL PGAPLAAENH401 FPETVLTLGD SHAGHLRGFL DYVGSREGWK AKILSLDSEC LVWVDEKLAD 451NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF LIPGFPARFR 501 ETVKRIAAVKPVYVFANNTS ISRSPLREEK LKRFAANQYL RPIQAMGDIG 551 KSNQAVFDLI KDIPNVHWVDAQKYLPKNTV EIYGRYLYGD QDHLTYFGSY 601 YMGREFHKHE RLLKSSRDGA LQ*

ORF128a and ORF128-1 show 99.5% identity in 622 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF128 shows 93.4% identity over 244 aa overlap with a predicted ORF(ORF128ng) from N. gonorrhoeae:

The complete length ORF128ng nucleotide sequence <SEQ ID 833> is:

1 ATGCAAGCTG TCCGATACAG GCCTGAAATT GACGGATTGC GGGCCGTCGC 51CGTGCTATCC GTCATTATTT TCCACCTGAA TAACCGCTGG CTGCCCGGAG 101GATTCCTGGG GGTGGACATT TTCTTTGTCA TCTCGGGATT CCTCATTACC 151AACATCATTC TTTCTGAAAT ACAGAACGGT TCTTTTTCTT TCCGGGATTT 201TTATACCCGC AGGATTAAGC GGATTTATCC TGCTTTTATT GCGGCCGTGT 251CCCTGGCTTC GGTGATTGCT TCTCAAATCT TCCTTTACGA AGATTTCAAC 301CAAATGAGGA AAACCATAGA GCTTTCTACG GTTTTTTTGT CCAATATTTA 351TTTGGGGTTC CGATTGGGGT ATTTCGATTT GAGTGCCGAC GAGAACCCCG 401TACTGCATAT CTGGTCTTTG GCGGTAGAGG AACAGTATTA CCTCCTGTAT 451CCTCTTTTGC TGATATTCTG TTACAAAAAA ACCAAATCAC TACGGGTGCT 501GCGTAATATC AGCATCATCC TGTTTCTGAT TTTGACCGCA TCATCGTTTT 551TGCCGGCCGG GTTTTATACC GACATCCTCA ACCAACCcaa TACTTATTAC 601CTTTCGACAC TGAGGTTTCC CGAGCTGTTG GTGGGTTCGC TGTTGGCGGT 651TTACGGGCAA ACGCAAAACG GCAGACGGCA AACAGAAAAT GGAAAACGGC 701AGTTGCTTTC ATTACTCTGT TTCGGCGCat tgCTTGTCTG CCTGTTCGTG 751ATCGACAAAC ACGATCCGTT TATCCCGGGA ATAACCCTGC TCCTTCCCTG 801CCTGCTGACG GCGCTGCTTA TCCGGAGTAT GCAATACGGG ACACTTCCGA 851CCCGCATCCT GTCGGCAAGC CCCATCGTAT TTGTCGGCAA AATCTCTTAT 901TCCCTATACC TGTACCATTG GATTTTTATT GCCTTCGCCC ATTACATTAC 951AGGCGACAAA CAGCTCGGAC TGCCTGCCGT ATCGGCGGTT GCCGCGTTGA 1001CGGCCGGATT TTCCCTGTTG AGCTATTATT TGATTGAACA GCCGCTTAGA 1051AAACGGAAGA TGACCTTCAA AAAGGCATTT TTCTGCCTTT ATCTCGCCCC 1101GTCCCTGATG CTTGTCGGTT ACAACCTGTA TTCAAGAGGG ATATTGAAAC 1151AGGAACACCT CCGCCCGCTG CCCGGCACGC CCGTTGCTGC GGAAAATAAT 1201TTTCCGGAAA CCGTCTTGAC CCTCGGCGAC TCGCACGCCG GACACCTGCG 1251GGGGTTTCTG GATTATGTCG GCGGCAGGGA AGGGTGGAAA GCTAAAATCC 1301TGTCCCTCGA TTCGGAGTGT TTGGTTTGGG TGGATGAGAA GCTGGCAGAC 1351AACCCGTTGT GCCGAAAATA CCGGGATGAA GTTGAAAAAG CCGAAGCTGT 1401TTTCATTGCC CAATTCTATG ATTTGAGGAT GGGCGGCCAG CCCGTGCCGA 1451GATTTGAAGC GCAATCCTTC CTGATACCCG GGTTCAAAGC CCGATTCAGG 1501GAAACCGTCA AGAGGATAGC CGCCGTCAAA CCTGTATATG TTTTTGCAAA 1551CAATACATCA ATCAGCCGTT CTCCCTTGAG GGAGGAAAAA TTGAAAAGAT 1601TTGCTATAAA CCAATACCTC CGGCCTATTC GGGCTATGGG CGACATCGGC 1651AAGAGCAATC AGGCGGTCTT TGATTTGGTT AAAGATATTC CCAATGTGCA 1701TTGGGTGGAC GCACAAAAAT ACCTGCCCAA AAACACGGTC GAAATACACG 1751GACGCTATCT TTACGGCGAC CAAGACCACC TGACCTATTT CGGTTCTTAT 1801TATATGGGGC GGGAATTTCA CAAACACGAA CGCCTGCTCA AGCATTCCCG 1851AGGCGGCGCA TTGCAGTAG

This encodes a protein having amino acid sequence <SEQ ID 834>:

1 MQAVRYRPEI DGLRAVAVLS VIIFHLNNRW LPGGFLGVDI FFVISGFLIT 51NIILSEIQNG SFSFRDFYTR RIKRIYPAFI AAVSLASVIA SQIFLYEDFN 101QMRKTIELST VFLSNIYLGF RLGYFDLSAD ENPVLHIWSL AVEEQYYLLY 151PLLLIFCYKK TKSLRVLRNI SIILFLILTA SSFLPAGFYT DILNQPNTYY 201LSTLRFPELL VGSLLAVYGQ TQNGRRQTEN GKRQLLSLLC FGALLVCLFV 251IDKHDPFIPG ITLLLPCLLT ALLIRSMQYG TLPTRILSAS PIVFVGKISY 301SLYLYHWIFI AFAHYITGDK QLGLPAVSAV AALTAGFSLL SYYLIEQPLR 351KRKMTFKKAF FCLYLAPSLM LVGYNLYSRG ILKQEHLRPL PGTPVAAENN 401FPETVLTLGD SHAGHLRGFL DYVGGREGWK AKILSLDSEC LVWVDEKLAD 451NPLCRKYRDE VEKAEAVFIA QFYDLRMGGQ PVPRFEAQSF LIPGFKARFR 501ETVKRIAAVK PVYVFANNTS ISRSPLREEK LKRFAINQYL RPIRAMGDIG 551KSNQAVFDLV KDIPNVHWVD AQKYLPKNTV EIHGRYLYGD QDHLTYFGSY 601YMGREFHKHE RLLKHSRGGA LQ*

ORF128ng and ORF128-1 show 95.7% identity in 622 aa overlap:

In addition, ORF218ng shows homology to a hypothetical H. influenzaeprotein:

sp|P43993|Y392_HAEIN HYPOTHETICAL PROTEIN HI0392 >gi|1074385|pir||B64007hypothetical protein HI0392 - Haemophilus influenzae (strain Rd KW20)>gi|1573364 (U32723) H. influenzae predicted coding region HI0392[Haemophilus influenzae] Length = 245 Score = 239 bits (604), Expect =3e−62 Identities = 124/225 (55%), Positives = 152/225 (67%), Gaps =1/225 (0%) Query: 38VDIFFVISGFLITNIILSEIQNGSFSFRDFYTRRIKRIYPXXXXXXXXXXXXXXXXFLYE 97+DIFFVISGFLIT II++EIQ  SFS + FYTRRIKRIYP                F+Y Sbjct: 1MDIFFVISGFLITGIIITEIQQNSFSLKQFYTRRIKRIYPAFITVMALVSFIASAIFIYN 60 Query:98 DFNQMRKTIELSTVFLSNIYLGFRLGYFDLSADENPVLHIWSLAVEEQXXXXXXXXXIFC 157DFN++RKTIEL+  FLSN YLG   GYFDLSA+ENPVLHIWSLAVE Q         I Sbjct: 61DFNKLRKTIELAIAFLSNFYLGLTQGYFDLSANENPVLHIWSLAVEGQYYLIFPLILILA 120 Query:158 YKKTKSLRVLRNISIILFLILTASSFLPAGFYTDILNQPNTYYLSTLRFPELLVGSLLAV 217YKK + ++VL  I++ILF IL A+SF+ A FY ++L+QPN YYLS LRFPELLVGSLLA+ Sbjct: 121YKKFREVKVLFIITLILFFILLATSFVSANFYKEVLHQPNIYYLSNLRFPELLVGSLLAI 180 Query:218 YGQTQNGRRQTENGKRQLLSLLCFGALLVCLFVIDKHDPFIPGIT 262 Y    N +Q       +L++L    L  CLF+++ +  FIPGIT Sbjct: 181YHNLSN-KVQLSKQVNNILAILSTLLLFSCLFLMNNNIAFIPGIT 224

This analysis, including the identification of several putativetransmembrane domains, suggests that these proteins from N. meningitidisand N. gonorrhoeae, and their epitopes, could be useful antigens forvaccines or diagnostics, or for raising antibodies.

Example 99

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 835>:

1 ..ATTATTTACG AATACCGCTG GATGTTTCTT TACGGCGCAC TGACGACCTT 51  GGGGCTGACG GTCGTGGCAA C.GCGGGCGG TTCGGTATTG GGTCTGTTGT 101  TGGCGTTGGC GCGCCTGATT CACTTGGAAA AAGCCGGTGC GCCGATGCGC 151  GTGCTGGCGT GGGCGTTGCG TAAAGTTTCG CTGCTGTATG TTACGCTGTT 201  CCGGGGTACG CCGCTGTTTG TGCAGATTGT GATTTGGGCG TATGTGTGGT 251  TTCCGTTTTT CGTC..

This corresponds to the amino acid sequence <SEQ ID 836; ORF129>:

1 ..IIYEYRWMFL YGALTTLGLT VVAXAGGSVL GLLLALARLI HLEKAGAPMR 51  VLAWALRKVS LLYVTLFRGT PLFVQIVIWA YVWFPFFV..

Further work revealed the complete nucleotide sequence <SEQ ID 837>:

1 ATGGATTTTC GTTTTGACAT TATTTACGAA TACCGCTGGA TGTTTCTTTA 51CGGCGCACTG ACGACCTTGG GGCTGACGGT CGTGGCAACG GCGGGCGGTT 101CGGTATTGGG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA CTTGGAAAAA 151GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA AAGTTTCGCT 201GCTGTATGTT ACGCTGTTCC GGGGTACGCC GCTGTTTGTG CAGATTGTGA 251TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC AGACGGCATT 301TTGGTCAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT ACGGGCCGCT 351GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG TATATCTGTG 401AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA GATGGAGGCG 451GCGCGTTCTT TGGGGCTGAC CTATCCGCAG GCGATGCGCT ATGTGATTCT 501GCCGCAGGCA TTGCGCCGCA TGCTGCCGCC TTTGGCGAGC GAGTTCATCA 551CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT GGCGGAGTTG 601GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT ATGAAGAACC 651GCTTTACACC GTCGCCCTGA TTTATCTGTT GATGACGACT TTCTTAGGCT 701GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA CCGCTGA

This corresponds to the amino acid sequence <SEQ ID 838; ORF129-1>:

1 MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL ALARLIHLEK 51AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF PFFV HPSDGI 101LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI QSIDKGQMEA 151ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS LLSVIAVAEL 201AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE KRYNPQHR*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF129 shows 98.9% identity over a 88aa overlap with an ORF (ORF129a)from strain A of N. meningitidis:

The complete length ORF129a nucleotide sequence <SEQ ID 839> is:

1 ATGGATTTTC GTTTTGACAT TATTTACGAA TACCGCTGGA TGTTTCTTTA 51CGGCGCACTG ACGACCTTGG GGCTGACGGT CGTGGCGACG GCGGGCGGTT 101CGGTATTGGG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA CTTGGAAAAA 151GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA AGGTTTCGCT 201GCTGTATGTT ACGCTGTTCC GGGGTACGCC GCTGTTTGTG CAGATTGTGA 251TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC AGACGGCATT 301TTGGTTAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT ACGGGCCGCT 351GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG TATATCTGTG 401AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA GATGGAGGCG 451GCGCGTTCTT TGGGGCTGAC CTATCCGCAG GCGATGCGCT ATGTGATTCT 501GCCGCAGGCA TTGCGCCGTA TGCTGCCGCC TTTGGCGAGC GAGTTCATCA 551CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT GGCGGAGTTG 601GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT ATGAAGAACC 651GCTTTACACC GTCGCCCTGA TTTATCTGTT GATGACGACT TTCTTAGGCT 701GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA CCGCTGA

This encodes a protein having amino acid sequence <SEQ ID 840>:

1 MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL ALARLIHLEK 51AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF PFFVHPSDGI 101LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI QSIDKGQMEA 151ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS LLSVIAVAEL 201AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE KRYNPQHR*

ORF129a and ORF129-1 show 100.0% identity in 248 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF129 shows 98.9% identity over a 88 aa overlap with a predicted ORF(ORF129ng) from N. gonorrhoeae:

An ORF129ng nucleotide sequence <SEQ ID 841> was predicted to encode aprotein having amino acid sequence <SEQ ID 842>:

1 MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL ALARLIHLEK 51AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF PFFVILHTAF 101LGNAMRQSRR VPDKGRWIAG SLELNCQPRG RKTRGEFPPG ESNLGTEPRN 151PLSMGQRRFP GCENWYPPQN FIKK*

Further work revealed the following gonococcal sequence <SEQ ID 843>:

1 ATGGATTTTc gtTTTGACAT TATTTAcgaA TACCGCTGGA TGTTTCTTTA 51CGGCGCACTG Acgaccttgg ggctgacggt cgtggcgacg gCGGGCGGTT 101CGGtattggG TCTGTTGTTG GCGTTGGCGC GCCTGATTCA CTTGGAAAAA 151GCCGGTGCGC CGATGCGCGT GCTGGCGTGG GCGTTGCGTA AGGTTTCGCT 201GCTGTACGTT ACCCTGTTCC GGGGTACGCC GCTGTTTGTG CAGATTGTGA 251TTTGGGCGTA TGTGTGGTTT CCGTTTTTCG TCCATCCTTC AGACGGCATT 301TTGGTCAGCG GCGAGGCGGC AATCGCGCTG CGTCGCGGAT ACGGGCCGCT 351GATTGCCGGT TCTTTGGCAC TGATCGCCAA CTCGGGGGCG TATATCTGTG 401AGATTTTCCG CGCGGGCATC CAGTCTATAG ACAAAGGACA GATGGAGGCG 451GCGTGTTCTT TGGGACTGAC CTATCCGCAG GCGATGCGCT ATGTGATTCT 501GCCGCAGGCA TTGCGCCGTA TGCTGCCGCC TTTGGCGAGC GAGTTCATCA 551CGCTCTTGAA AGACAGCTCG CTGCTGTCGG TCATTGCTGT GGCGGAGTTG 601GCGTATGTTC AGAATACGAT TACGGGCCGG TATTCGGTTT ATGAAGAACC 651GCTTTACACC GCCGCCCTGA TTTATCTGTT GATGACGACT TTCTTAGGCT 701GGATATTCCT GCGTTTGGAA AAACGTTACA ATCCGCAACA CCGCTGA

This corresponds to the amino acid sequence <SEQ ID 844; ORF129ng-1>:

1 MDFRFDIIYE YRWMFLYGAL TTLGLTVVAT AGGSVLGLLL ALARLIHLEK 51AGAPMRVLAW ALRKVSLLYV TLFRGTPLFV QIVIWAYVWF PFFVHPSDGI 101LVSGEAAIAL RRGYGPLIAG SLALIANSGA YICEIFRAGI QSIDKGQMEA 151ARSLGLTYPQ AMRYVILPQA LRRMLPPLAS EFITLLKDSS LLSVIAVAEL 201AYVQNTITGR YSVYEEPLYT VALIYLLMTT FLGWIFLRLE KRYNPQHR*

ORF129ng-1 and ORF129-1 show 99.2% identity in 248 aa overlap:

In addition, ORF129ng-1 is homologous to an ABC transporter from A.fulgidus:

2650409(AE001090) glutamine ABC transporter, permease protein (glnP)[Archaeoglobus fulgidus] Length = 224 Score = 132 bits (329), Expect =2e−30 Identities = 86/178 (48%), Positives = 103/178 (57%), Gaps =18/178 (10%) Query: 65VSLLYVTLFRGTPLFVQIVIWAYVWFPFFVHPSDGILVSGEAAIALRRGYGPLIAGSLAL 124+S  YV + RGTPL VQI+I       +F  P+ GI +  E A            G +AL Sbjct: 58ISTAYVEVIRGTPLLVQILI------VYFGLPAIGINLQPEPA------------GIIAL 99 Query:125 IANSGAYICEIFRAGIQSIDKGQMEAACSLGLTYPQAMRYVILPQALRRMLPPLASEFIT 184   SGAYI EI RAGI+SI  GQMEAA SLG+TY QAMRYVI PQA R +LP L +EFI Sbjct: 100SICSGAYIAEIVRAGIESIPIGQMEAARSLGMTYLQAMRYVIFPQAFRNILPALGNEFIA 159 Query:185 LLKDSSLLSVIAVAELAYVQNTITGRYSVYEEPLYTAALIYLLMTTFLGWIFLRLEKR 242LLKDSSLLSVI++ EL  V   I         P    AL YL+MT  L  +    +K+ Sbjct: 160LLKDSSLLSVISIVELTRVGRQIVNTTFNAWTPFLGVALFYLMMTIPLSRLVAYSQKK 217

This analysis, including the identification of transmembrane domains inthe two proteins, suggests that the proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

Example 100

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 845>:

1 ..CTGAAAGAAT GCCGTCTGAA AGACCCTGTT TTTATTCCAA ATATCGTTTA 51  TAAGAACATC GCCATTACTT TCCTGCTCTT GCACGCCGCC GCCGAACTTT 101  GGCTGCCCGC GCAAACCGCC GGTTTTACCG CGCTCGCCGT CGGCTTCATC 151  CTGCTCGCCA AGCTGCGTGA gCTTCACCAT CACGAACTCT TACGTAAACA 201  cTACGTCCGC ACTTATTACy TGCTCCAACT CTTTGCCGCC GCAGgcTAgT 251  TTGTGGACAG GCGCGGCGwA ATTACAAAAC CTGCCCGCyT CCGCGCCCCT 301  GCACCTGATT ACCCTCGGCG GCATGATGGG CGGCGTGATG ATGGTGTGGc 351  TGACCGCCGG ACTGTGGCAC AGCGGCTTTA CCAAACTCGA CTACCCCAAA 401  CTCTGCCGCA TTGCCGTCCC CATCCTTTTC GCCGCCGCCG TCTCGCGCGC 451  TTTCTTGrTG AACGTGAACC CGrTATTTTT CATTACCGTT CCTGCGATTC 501  TGACCGCCGC CGTATTCGTA CTGTATCTTT TCrCGTTTAT ACCGATATTT 551  CGGGCGAATG CGTTTACAGA CGATCCGGAr TAr

This corresponds to the amino acid sequence <SEQ ID 846; ORF130>:

1 ..LKECRLKDPV FIPNIVYKNI AITFLLLHAA AELWLPAQTA GFTALAVGFI 51  LLAKLRELHH HELLRKHYVR TYYLLQLFAA AGSLWTGAAX LQNLPASAPL 101  HLITLGGMMG GVMMVWLTAG LWHSGFTKLD YPKLCRIAVP ILFAAAVSRA 151  FLXNVNPXFF ITVPAILTAA VFVLYLFXFI PIFRANAFTD DPE*

Further work revealed the complete nucleotide sequence <SEQ ID 847>:

1 ATGCGGCCGT TTTTCGTCGG CGCGGCGGTG CTTGCCATAC TCGGTGCGCT 51GGTGTTTTTC ATCAACCCCG GTGCCATCGT CCTGCACCGC CAAATTTTCT 101TGGAACTTAT GCTGCCGGCG GCATACGGCG GTTTTTTGAC TGCGGCTTTG 151TTGGACTGGA CGGGTTTTTC GGGTAACCTG AAACCTGTCG CGACTTTGAT 201GGCGGCATTA TTGCTCGCCG CATCCGCTAT ACTGCCCTTT TCGCCGCAAA 251CTGCCTCGTT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT GCTGTTCTGC 301GCCCGGCTGA TTTGGCTAGA CCGAAACACC GACAACTTCG CCCTGCTAAT 351GTTACTTGCC GCGTTCACTG TTTTTCAGAC GGCATATGCC GTCAGCGGCG 401ATTTGAACCT GTTGCGCGCG CAAGTGCATC TAAATATGGC GGCGGTGATG 451TTCGTATCCG TGCGCGTCAG TATTCTTTTG GGCGCGGAAG CCCTGAAAGA 501ATGCCGTCTG AAAGACCCTG TTTTTATTCC AAATATCGTT TATAAAAACA 551TCGCCATTAC TTTCCTGCTC TTGCACGCCG CCGCCGAACT TTGGCTGCCC 601GCGCAAACCG CCGGTTTTAC CGCGCTCGCC GTCGGCTTCA TCCTGCTCGC 651CAAGCTGCGT GAGCTTCACC ATCACGAACT CTTACGTAAA CACTACGTCC 701GCACTTATTA CCTGCTCCAA CTCTTTGCCG CCGCAGGCTA TTTGTGGACA 751GGCGCGGCGA AATTACAAAA CCTGCCCGCC TCCGCGCCCC TGCACCTGAT 801TACCCTCGGC GGCATGATGG GCGGCGTGAT GATGGTGTGG CTGACCGCCG 851GACTGTGGCA CAGCGGCTTT ACCAAACTCG ACTACCCCAA ACTCTGCCGC 901ATTGCCGTCC CCATCCTTTT CGCCGCCGCC GTCTCGCGCG CTTTCTTGAT 951GAACGTGAAC CCGATATTTT TCATTACCGT TCCTGCGATT CTGACCGCCG 1001CCGTATTCGT ACTGTATCTT TTCACGTTTA TACCGATATT TCGGGCGAAT 1051GCGTTTACAG ACGATCCGGA ATAA

This corresponds to the amino acid sequence <SEQ ID 848; ORF130-1>:

1 MRPFFVGAAV LAILGALVFF INPGAIVLHR QIFLELMLPA AYGGFLTAAL 51LDWTGFSGNL KPVATLMAAL LLAASAILPF SPQTASFFVA AYWLVLLLFC 101ARLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA QVHLNMAAVM 151FVSVRVSILL GAEALKECRL KDPVFIPNIV YKNIAITFLL LHAAAELWLP 201AQTAGFTALA VGFILLAKLR ELHHHELLRK HYVRTYYLLQ LFAAAGYLWT 251GAAKLQNLPA SAPLHLITLG GMMGGVMMVW LTAGLWHSGF TKLDYPKLCR 301IAVPILFAAA VSRAFLMNVN PIFFITVPAI LTAAVFVLYL FTFIPIFRAN 351 AFTDDPE*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF130 shows 94.3% identity over a 193aa overlap with an ORF (ORF130a)from strain A of N. meningitidis:

The complete length ORF130a nucleotide sequence <SEQ ID 849> is:

1 ATGCGGCCGT TTTTCGTCGG CGCGGCGGTG CTTGCCATAC TCGGTGCGCT 51GGTGTTTTTC ATCAACCCCG GTGCCATCGT CCTGCACCGC CAAATTTTCT 101TGGAACTTAT GCTGCCGGCG GCATACGGCG GTTTTTTGAC TGCGGCTTTG 151TTGGACTGGA CGGGTTTTTC GGGTAACCTG AAACCTGTCG CGACTTTGAT 201GGCGGCATTA TTGCTCGCCG CATCCGCTAT ACTGCCCTTT TCGCCGCAAA 251CTGCCTCGTT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT GCTGTTCTGC 301GCCCGGCTGA TTTGGCTAGA CCGAAACACC GACAACTTCG CCCTGCTAAT 351GTTACTTGCC GCGTTCACTG TTTTTCAGAC GGCATATGCC GTCAGCGGCG 401ATTTGAACCT GTTGCGCGCG CAAGTGCATC TAAATATGGC GGCGGTGATG 451TTCGTATCCG TGCGCGTCAG TATTCTTTTG GGCGCGGAAG CCCTGAAAGA 501ATGCCGTCTG AAAGACCCAG TATTCATCCC CAATGTCGTC TATAAAAACA 551TCGCCATTAC CTTCCTGCTC CTGCACGCCG CCGCCGAACT TTGGCTGCCT 601GCGCAAACCG CCGGTTTTAC CTCGCTCGCC GTCGGCTTTA TCCTGCTTGC 651CAAGCTGCGT GAGCTTCACC ATCACGAACT CCTGCGCAAA CACTACGTCC 701GCACTTATTA CCTGCTCCAA CTCTTTGCCG CCGCAGGCTA TTTGTGGACA 751GGCGCGGCGA AATTACAAAA CCTGCCCGCC TCCGCGCCCC TGCACCTGAT 801TACCCTCGGT GGCATGATGG GCAGCGTGAT GATGGTGTGG CTGACTGCCG 851GACTGTGGCA CAGCGGCTTT ACCAAGCTCG ACTACCCGAA ACTCTGCCGC 901ATCGCCGTCC CCATCCTNTT CGCCGCCGCC GTTTCGCGCG CTGTTTTAAT 951GAACGTAAAC CCGATATTCT TCATCACCGT CCCCGCAATT CTGACCGCCG 1001CCGTGTTCGT GCTTTACCTG CTGACATTCG TACCGATCTT TCGGGCGAAC 1051GCGTTTACAG ACGATCCGGA ATAA

This encodes a protein having amino acid sequence <SEQ ID 850>:

1 MRPFFVGAAV LAILGALVFF INPGAIVLHR QIFLELMLPA AYGGFLTAAL 51LDWTGFSGNL KPVATLMAAL LLAASAILPF SPQTASFFVA AYWLVLLLFC 101ARLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA QVHLNMAAVM 151FVSVRVSILL GAEALKECRL KDPVFIPNVV YKNIAITFLL LHAAAELWLP 201AQTAGFTSLA VGFILLAKLR ELHHHELLRK HYVRTYYLLQ LFAAAGYLWT 251GAAKLQNLPA SAPLHLITLG GMMGSVMMVW LTAGLWHSGF TKLDYPKLCR 301IAVPILFAAA VSRAVLMNVN PIFFITVPAI LTAAVFVLYL LTFVPIFRAN 351 AFTDDPE*

ORF130a and ORF130-1 show 98.3% identity in 357 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF130 shows 91.7% identity over a 193 aa overlap with a predicted ORF(ORF130ng) from N. gonorrhoeae:

An ORF130ng nucleotide sequence <SEQ ID 851> was predicted to encode aprotein having amino acid sequence <SEQ ID 852>:

1 MNKFFTHPMR PFFVGAAVLA ILGALVFFHQ PRRYHPAPPN FLGTYAAGCI 51RRFFDYRFVG PDGFFRQPET CRYFDGGVVA CCGCFIAVFT ATCRIFRRRL 101LAGVAAVLRL ADLARRQHRT LRSVDVTAAF TVFQTAYAVS GDLNLLRAQV 151HLNMAAVMFV SVRVSVLLGT ETLKECRLKD PVFIPNVIYK NIAITLLLHA 201AAELWLPAQT AGFTALAVGF ILLAKLRELH HHELLRKHYV RTYYLLQLFA 251AAGYLWTGAA KLQNLPASAP LHLITLGGMT GGVMMVWLTA GLWHSGFTKL 301DYPKLCRIAV SILFASAVSR AVLMNVNPIF FITVPEILTA AVFMLYLLTF 351VPIFRANAFT DDPE*

Further work revealed the following gonococcal DNA sequence <SEQ ID853>:

1 ATGCGCCCGT TTTTCGTCGG TGCGGCAGTA CTTGCCATAC TCGGTGCGTT 51GGTGTTTTTT ATCAACCCCG GCGCTATCAT CCTGCACCGC CAAATTTTCT 101TGGAACTTAT GCTGCCGGCT GCATACGGCG GTTTTTTGAC TACCGCTTTG 151TTGGACCGGA CGGGTTTTTC AGGCAACCTG AAACCTGCCG CTACTTTGAT 201GGCGGTGTTG TTGCTTGTTG CGGCTGTTTT ATTGCCGTTT TTACCGCAAC 251TTGCCGCATT TTTCGTCGCC GCCTATTGGC TGGTGTTGCT GCTGTTCTGC 301GCCTGGCTGA TTTGGCTCGA CCGCAACACC GACAACTTCG CTCTGTTGAT 351GTTACTTGCC GCATTTACCG TTTTTCAGAC GGCCTATGCC GTCAGCGGCG 401ATTTGAACTT ACTGCGCGCG CAAGTGCATT TGAATATGGC GGCGGTCATG 451TTCGTATCCG TCCGCGTCAG CGTCCTTTTG GGCACGGAAA CCCTGAAAGA 501ATGCCGTCTG AAAGACCCCG TATTCATCCC CAACGTTATC TATAAAAACA 551TCGCCATCAC CCTGCTGCTG CACGCCGCCG CCGAACTTTG GCTGCCCGCG 601CAAACCGCCG GTTTTACTGC GCTTGCCGTC GGCTTCATCC TGCTCGCCAA 651GCTGCGCGAA CTGCACCATC ACGAACTCTT ACGCAAACAC TACGTCCGCA 701CTTATTACCT GCTCCAGCTC TTTGCCGCCG CAGGTTATCT GTGGACAGGC 751GCGGCGAAAC TGCAAAACCT GCCCGCCTCC GCGCCCCTGC ACCTGATTAC 801CCTCGGCGGC ATGACGGGTG GCGTGATGAT GGTGTGGCTG ACTGCCGGAC 851TGTGGCACAG CGGCTTTACC AAACTCGACT ACCCGAAACT CTGCCGCATC 901GCCGTCTCCA TCCTTTTCGC CTCCGCCGTT TCGCGCGCTG TTTTAATGAA 951CGTGAATCCG ATATTCTTCA TCACCGTTCC CGAGATTCTG ACCGCCGCCG 1001TGTTCATGCT TTACCTGCTG ACGTTCGTAC CGATTTTTCG AGCGAACGCG 1051TTTACAGACG ATCCGGAATA A

This corresponds to the amino acid sequence <SEQ ID 854; ORF130ng-1>:

1 MRPFFVGAAV LAILGALVFF INPGAIILHR QIFLELMLPA AYGGFLTTAL 51LDRTGFSGNL KPAATLMAVL LLVAAVLLPF LPQLAAFFVA AYWLVLLLFC 101AWLIWLDRNT DNFALLMLLA AFTVFQTAYA VSGDLNLLRA QVHLNMAAVM 151FVSVRVSVLL GTETLKECRL KDPVFIPNVI YKNIAITLLL HAAAELWLPA 201QTAGFTALAV GFILLAKLRE LHHHELLRKH YVRTYYLLQL FAAAGYLWTG 251AAKLQNLPAS APLHLITLGG MTGGVMMVWL TAGLWHSGFT KLDYPKLCRI 301AVSILFASAV SRAVLMNVNP IFFITVPEIL TAAVFMLYLL TFVPIFRANA 351 FTDDPE*

ORF130ng-1 and ORF130-1 show 92.4% identity in 357 aa overlap:

Based on this analysis, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 101

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 855>:

1 ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT TGCTTGCATT 51TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT TCGTCCCTCA 101CCGGCTGGTG TAAGCCGAGA AAACCGGCTG CCATCGATTT TTGGGATATT 151GGCGGCGAGA GTCCGCCGTC TTTAGGGGAC TACGAGATAC CGCTTTCAGA 201CGGCAATAGT TCCGTCAGGG CAAACGAATA TGAATCCGCA CAACAATCTT 251ACTTTTACAG GAAAATAGGG AAGTTTGAAG C.TGCGGGCT GGATTGGCGT 301ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG GAGGATTTGA 351CTGCTTGGAA AAG..

This corresponds to the amino acid sequence <SEQ ID 856; ORF131>:

1 MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLTGWCKPR KPAAIDFWDI 51GGESPPSLGD YEIPLSDGNS SVRANEYESA QQSYFYRKIG KFEXCGLDWR 101TRDGKPLIET FKQGGFDCLE K..

Further work revealed the complete nucleotide sequence <SEQ ID 857>:

1 ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT TGCTTGCATT 51TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT TCGTCCCTCA 101CCGGCTGGTG TAAGCCGAGA AAACCGGCTG CCATCGATTT TTGGGATATT 151GGCGGCGAGA GTCCGCCGTC TTTAGGGGAC TACGAGATAC CGCTTTCAGA 201CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCA CAACAATCTT 251ACTTTTACAG GAAAATAGGG AAGTTTGAAG CCTGCGGGCT GGATTGGCGT 301ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG GAGGATTTGA 351CTGCTTGGAA AAGCAGGGGT TGCGGCGCAA CGGTCTGTCC GAGCGCGTCC 401 GATGGTAA

This corresponds to the amino acid sequence <SEQ ID 858; ORF131-1>:

1 MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLTGWCKPR KPAAIDFWDI 51GGESPPSLGD YEIPLSDGNR SVRANEYESA QQSYFYRKIG KFEACGLDWR 101TRDGKPLIET FKQGGFDCLE KQGLRRNGLS ERVRW*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF131 shows 95.0% identity over a 121 aa overlap with an ORF (ORF131a)from strain A of N. meningitidis:

The complete length ORF131a nucleotide sequence <SEQ ID 859> is:

1 ATGGAAATTC GGGCAATAAA ATATACGGCA ATGGCTGCGT TGCTTGCATT 51TACGGTTGCA GGCTGCCGGT TGGCAGGTTG GTATGAGTGT TCGTCCCTGT 101CCGGCTGGTG TAAGCCGAGA AAACCTGCCG CCATCGATTT TTGGGATATT 151GGCGGCGAGA GTCCTCCGTC TTTAGAGGAC TACGAGATAC CGCTTTCAGA 201CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCA CAACAATCTT 251ACTTTTACAG GAAAATAGGG AAGTTTGAAG CCTGCGGGTT GGATTGGCGT 301ACGCGTGACG GCAAACCTTT GATTGAGACG TTCAAACAGG AAGGTTTTGA 351TTGTTTGAAA AAGCAGGGGT TGCGGCGCAA CGGTCTGTCC GAGCGCGTCC 401 GATGGTAA

This encodes a protein having amino acid sequence <SEQ ID 860>:

1 MEIRAIKYTA MAALLAFTVA GCRLAGWYEC SSLSGWCKPR KPAAIDFWDI 51GGESPPSLED YEIPLSDGNR SVRANEYESA QQSYFYRKIG KFEACGLDWR 101TRDGKPLIET FKQEGFDCLK KQGLRRNGLS ERVRW*

ORF131a and ORF131-1 show 97.0% identity in 135 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF131 shows 89.3% identity over 121 aa overlap with a predicted ORF(ORF131ng) from N. gonorrhoeae:

A complete length ORF131ng nucleotide sequence <SEQ ID 861> waspredicted to encode a protein having amino acid sequence <SEQ ID 862>:

1 MEIRVIKYTA TAALFAFTVA GCRLAGWYEC LSLSGWCKPR KPAAIDFWDI 51GGESPLSLED YEIPLSDGNR SVRANEYESA QKSYFYRKIG KFEACGLDWR 101TRDGKPLVER FKQEGFDCLE KQGLRRNGLS ERVRW*

Further work revealed the following gonococcal DNA sequence <SEQ ID863>:

1 ATGGAAATTC GGGTAATAAA ATATACGGCA ACGGCTGCGT TGTTTGCATT 51TACGGTTGCA GGCTGCCGGC TGGCGGGGTG GTATGAGTGT TCGTCCTTGT 101CCGGCTGGTG TAAGCCGAGA AAACCTGCCG CCATCGATTT TTGGGATATT 151GGCGGCGAGA GtccgctGTC TTTAGAGGAC TACGAGATAC CGCTTTCAGA 201CGGCAATCGT TCCGTCAGGG CAAACGAATA TGAATCCGCG CAAAAATCTT 251ACTTTTATAG GAAAATAGGG AAGTTTGAAG CCTGCGGGTT GGATTGGCGT 301ACGCGTGACG GCAAACCTTT GGTTGAGAGG TTCAAACAGG AAGGTTTCGA 351CTGTTTGGAA AAGCAGGGGT TGCGGCGCAA CGGCCTGTCC GAGCGCGTCC 401 GATGGTAA

This corresponds to the amino acid sequence <SEQ ID 864; ORF131ng-1>:

  1 MEIRVIKYTA TAALFAFTVA GCRLAGWYEC SSLSGWCKPR KPAAIDFWDI  51GGESPLSLED YEIPLSDGNR SVRANEYESA QKSYFYRKIG KFEACGLDWR 101 TRDGKPLVERFKQEGFDCLE KQGLRRNGLS ERVRW*

ORF131ng-1 and ORF131-1 show 92.6% identity in 135 aa overlap:

Based on the presence of a predicted prokaryotic membrane lipoproteinlipid attachment site, it is predicted that the proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 102

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 865>

  1 ATGAAACACA TCCATATTAT CGGTATCGGC GGCACGTTTA TGGGCGGGCT  51TGCCGCCATT GCCAAAGAAG CGGGGTTTGA AGTCAGCGGT TGCGACGCGA 101 AGATGTATCCGCCGATGAGC ACCCAGCTCG AAGCCTTGGG TATAGACGTG 151 TATGAAGGCT TCGATGCCGCTCAGTTGGAC GAATTTAAAG CCGACGTTTA 201 CGTTATCGGC AATGTCGCCA AGCGCGGGATGGATGTGGTT GAAGCGATTT 251 TGAACCTCGG CCTGCCtTAT ATtTcCGGCC CGCAATGGCTGTCGGAAAAC 301 GTGCTGCACC ATCATTGGGT ACTCGGTGTG GCGGGGACgC ACGGCAAAAC351 GACCACCGCC TCCATGCTCG CATGGGTCTT GGAATATgCC GGCCTCGCGC 401CGGGCTTCCT TATtGGCGGC GTACC.GGAA AATttCGGCG TTTCCGCCCG 451 CCTGCCGCAAACGCCGCGCC AAGACCCGAA CAGCCAATCG CCGTTTTTcG 501 TCATCGAAGC CGACGAATACGACACCGCCT TTtTCGACAA ACGTTCTAAA 551 TtCGTGCATT ACCGTCCGCG TACCGCCGTGTTGAACAATC TGGAATTCGA 601 CCACGCCGAC ATCTTTGCCG ACTTGGGCGC GATACAGACcCAGTTCCACT 651 ACCTCGTGCG TACCGTGCCG TCTGAAGGCT TAATCGTCTG CAACGGACGG701 CAGCAAAGCC TGCAAGATAC TTTGGACAAA GGCTGCTGGA CGCCGGTGGA 751AAAATTCGGC ACGGAACACG GCTGGCA..

This corresponds to the amino acid sequence <SEQ ID 866; ORF132>:

  1 MKHIHIIGIG GTFMGGLAAI AKEAGFEVSG CDAKMYPPMS TQLEALGIDV  51YEGFDAAQLD EFKADVYVIG NVAKRGMDVV EAILNLGLPY ISGPQWLSEN 101 VLHHHWVLGVAGTHGKTTTA SMLAWVLEYA GLAPGFLIGG VXGKFRRFRP 151 PAANAAPRPE QPIAVFRHRSRRIRHRLFRQ TFXIRALPSA YRRVEQSGIR 201 PRRHLCRLGR DTDPVPLPRA YRAVXRLNRLQRTAAKPARY FGQRLLDAGG 251 KIRHGTRLA..

Further work revealed the complete nucleotide sequence <SEQ ID 867>:

1 ATGAAACACA TCCATATTAT CGGTATCGGC GGCACGTTTA TGGGCGGGCT 51 TGCCGCCATTGCCAAAGAAG CGGGGTTTGA AGTCAGCGGT TGCGACGCGA 101 AGATGTATCC GCCGATGAGCACCCAGCTCG AAGCCTTGGG TATAGACGTG 151 TATGAAGGCT TCGATGCCGC TCAGTTGGACGAATTTAAAG CCGACGTTTA 201 CGTTATCGGC AATGTCGCCA AGCGCGGGAT GGATGTGGTTGAAGCGATTT 251 TGAACCTCGG CCTGCCTTAT ATTTCCGGCC CGCAATGGCT GTCGGAAAAC301 GTGCTGCACC ATCATTGGGT ACTCGGTGTG GCGGGGACGC ACGGCAAAAC 351GACCACCGCC TCCATGCTCG CATGGGTCTT GGAATATGCC GGCCTCGCGC 401 CGGGCTTCCTTATTGGCGGC GTACCGGAAA ATTTCGGCGT TTCCGCCCGC 451 CTGCCGCAAA CGCCGCGCCAAGACCCGAAC AGCCAATCGC CGTTTTTCGT 501 CATCGAAGCC GACGAATACG ACACCGCCTTTTTCGACAAA CGTTCTAAAT 551 TCGTGCATTA CCGTCCGCGT ACCGCCGTGT TGAACAATCTGGAATTCGAC 601 CACGCCGACA TCTTTGCCGA CTTGGGCGCG ATACAGACCC AGTTCCACTA651 CCTCGTGCGT ACCGTGCCGT CTGAAGGCTT AATCGTCTGC AACGGACGGC 701AGCAAAGCCT GCAAGATACT TTGGACAAAG GCTGCTGGAC GCCGGTGGAA 751 AAATTCGGCACGGAACACGG CTGGCAGGCC GGCGAAGCCA ATGCCGACGG 801 CTCGTTCGAC GTGTTGCTCGACGGCAAAAC CGCCGGACGC GTCAAATGGG 851 ATTTGATGGG CAGGCACAAC CGCATGAACGCGCTCGCCGT CATTGCCGCC 901 GCGCGTCATG TCGGTGTCGA TATTCAGACC GCCTGCGAAGCCTTGGGCGC 951 GTTTAAAAAC GTCAAACGCC GGATGGAAAT CAAAGGCACG GCAAACGGCA1001 TCACCGTTTA CGACGACTTC GCCCACCACC CGACCGCCAT CGAAACCACG 1051ATTCAAGGTT TGCGCCAACG CGTCGGCGGC GCGCGCATCC TCGCCGTCCT 1101 CGAACCGCGTTCCAACACGA TGAAGCTGGG CACGATGAAG TCCGCCCTGC 1151 CTGTAAGCCT CAAAGAAGCCGACCAAGTGT TCTGCTACGC CGGCGGCGTG 1201 GACTGGGACG TCGCCGAAGC CCTCGCGCCTTTGGGCGGCA GGCTGAACGT 1251 CGGCAAAGAC TTCGATGCCT TCGTTGCCGA AATCGTGAAAAACGCCGAAG 1301 TAGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG CGGAATACAC1351 GGAAAGCTGC TGGAAGCTTT GAGATAG

This corresponds to the amino acid sequence <SEQ ID 868; ORF132-1>:

  1 MKHIHIIGIG GTFMGGLAAI AKEAGFEVSG CDAKMYPPMS TQLEALGIDV  51YEGFDAAQLD EFKADVYVIG NVAKRGMDVV EAILNLGLPY ISGPQWLSEN 101 VLHHHWVLGVAGTHGKTTTA SMLAWVLEYA GLAPGFLIGG VPENFGVSAR 151 LPQTPRQDPN SQSPFFVIEADEYDTAFFDK RSKFVHYRPR TAVLNNLEFD 201 HADIFADLGA IQTQFHYLVR TVPSEGLIVCNGRQQSLQDT LDKGCWTPVE 251 KFGTEHGWQA GEANADGSFD VLLDGKTAGR VKWDLMGRHNRMNALAVIAA 301 ARHVGVDIQT ACEALGAFKN VKRRMEIKGT ANGITVYDDF AHHPTAIETT351 IQGLRQRVGG ARILAVLEPR SNTMKLGTMK SALPVSLKEA DQVFCYAGGV 401DWDVAEALAP LGGRLNVGKD FDAFVAEIVK NAEVGDHILV MSNGGFGGIH 451 GKLLEALR*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Hypothetical o457 Protein of E. coli (Accession NumberU14003)

ORF132 and o457 show 58% aa identity in 140 aa overlap:

Orf132: 4 IHIIGIGGTFMGGLAAIAKEAGFEVSGCDAKMYPPMSTQLEALGIDVYEGFDAAQLDEFK 63 IHI+GI GTFMGGLA +A++ G EV+G DA +YPPMST LE  GI++ +G+DA+QL+  + o457: 3IHILGICGTFMGGLAMLARQLGHEVTGSDANVYPPMSTLLEKQGIELIQGYDASQLEP-Q  61 Orf132:64 ADVYVIGNVAKRGMDVVEAILNLGLPYISGPQWLSENVLHHHWVLGVAGTHGKTTTASML 123 D+ +IGN   RG   VEA+L   +PY+SGPQWL + VL   WVL VAGTHGKTTTA M o457: 62PDLVIIGNAMTRGNPCVEAVLEKNIPYMSGPQWLHDFVLRDRWVLAVAGTHGKTTTAGMA 121 Orf132:124 AWVLEYAGLAPGFLIGGVXG 143  W+LE  G  PGF+IGGV G o457: 122TWILEQCGYKPGFVIGGVPG 141Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF132 shows 74.6% identity over a 189aa overlap with an ORF (ORF132a)from strain A of N. meningitidis:

The complete length ORF132a nucleotide sequence <SEQ ID 869> is:

   1 ATGAAACACA TCCACATTAT CGGTATCGGC GGCACGTTTA TGGGTGGGAT  51 TGCCGCCATT GCCAAAGAAG CAGGGTTTGA ANTCAGCGGT TGCGATGCGA 101 AGATGTATCC GCCGATGAGC ACCCAGCTCG AAGCCTTGGG CATAGGCGTG 151 TATGAAGGCT TCGACACCGC GCAGTTGGAC GAATTTAAAG CCGACGTTTA 201 CGTTATCGGC AATGTCGCCA AGCGCGGGAT GGATGTGGTT GAAGCGATTT 251 TGAACCGTGG GCTGCCTTAT ATTTCCGGCC CGCAATGGCT GGCTGAAAAC 301 NTGCTGCACC ATCATTGGNN ACTCGGCGTG GCGGNGACGC ACGGCAAAAC 351 GACCACCGCG TCTATGCTCG CGTGGGTTTT GGAATATGCC GGACTCGCAC 401 CGGGCTTCNT TATCGGCGGC GTACCGGAAA ACTTCAGCGT TTCCGCCCGC 451 CTGCCGCAAA CGCCGCGCCA AGACCCGAAC AGCCAATCGC CGTTTTTCGT 501 CATTGAAGCC GACGAATACG ACACCGCGTT TTTCGACAAA CGCTCCAAAT 551 TCGTGCATTA CCGTCCGCGT ACCGCCGTGT TGAACAATCT GGAATTCGAC 601 CACGCCGACA TCTTCGCCGA TTTGGGCGCG ATACAGACCC AGTTCCACCA 651 CCTCGTGCGT ACCGTGCCGT CTGAAGGCCT CATCGTCTGC AACGGACGGC 701 AGCAAAGCCT GCAAGACACT TTGGACAAAG GCTGCTGGAC GCCGGTGGAA 751 AAATTCGGCA CGGAACACGG CTGGCAGGCC GGCGAAGCCA ATGCCGATGG 801 CTCGTTCGAC GTGTTGCTTG ACGGCAAAAA AGCCGGACAC GTCGCTTGGA 851 GTTTGATGGG CGGACACAAC CGCATGAACG CGCTCGCNGT CATCGCCGCC 901 GCGCGTCATG CCGGAGTNGA CATTCAGACG GCCTGCGAAG CCTTGAGCAC 951 GTTTAAAAAC GTCAAACGCC GCATGGAAAT CAAAGGCACG GCAAACGGTA1001 TCACCGTTTA CGACGACTTC GCCCACCATC CGACCGCTAT CGAAACCACG1051 ATTCAAGGTT TGCGCCAGCG CGTCGGCGGC GCGCGCATCC TCGCCGTCCT1101 CGAACCGCGT TCCAATACGA TGAAGCTGGG TACGATGAAA GCCGCCCTGC1151 CCGCAAGCCT CAAAGAAGCC GACCAAGTGT TCTGNTACGC CGGCGGCGCG1201 GACTGGGACG TTGCCGAAGC CCTCGCGCCT TTGGGCGGCA GGCTGCACGT1251 CGGCAAAGAC TTCGATGCCT TCGTTGCCGA AATCGTGAAA AACGCCGAAG1301 CAGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG CGGAATACAC1351 ACCAAACTGC TGGACGCTTT GAGATAG

This encodes a protein having-amino acid sequence <SEQ ID 870>:

  1 MKHIHIIGIG GTFMGGIAAI AKEAGFEXSG CDAKMYPPMS TQLEALGIGV 51 YEGFDTAQLD EFKADVYVIG NVAKRGMDVV EAILNRGLPY ISGPQWLAEN101 XLHHHWXLGV AXTHGKTTTA SMLAWVLEYA GLAPGFXIGG VPENFSVSAR151 LPQTPRQDPN SQSPFFVIEA DEYDTAFFDK RSKFVHYRPR TAVLNNLEFD201 HADIFADLGA IQTQFHHLVR TVPSEGLIVC NGRQQSLQDT LDKGCWTPVE251 KFGTEHGWQA GEANADGSFD VLLDGKKAGH VAWSLMGGHN RMNALAVIAA301 ARHAGVDIQT ACEALSTFKN VKRRMEIKGT ANGITVYDDF AHHPTAIETT351 IQGLRQRVGG ARILAVLEPR SNTMKLGTMK AALPASLKEA DQVFXYAGGA401 DWDVAEALAP LGGRLHVGKD FDAFVAEIVK NAEAGDHILV MSNGGFGGIH 451 TKLLDALR*

ORF132a and ORF132-1 show 93.9% identity in 458 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF132 shows 89.6% identity over 259 aa overlap with a predicted ORF(ORF132ng) from N. gonorrhoeae:

An ORF132ng nucleotide sequence <SEQ ID 871> was predicted to encode aprotein having amino acid sequence <SEQ ID 872>:

  1 MKHIHIIGIG GTFMGGIAAI AKEAGFKVSG CDAKMYPPMS TQLEALGIGV  51HEGFDAAQLE EFQADIYVIG NVARRGMDVV EAILNRGLPY ISGPQWLAEN 101 VLHHHWVLGVAGTHGKTTTA SMLAWVLEYA GLAPGFLIGG VPGKFRRFRP 151 PTANAASRPE QQIAVFRHRSRRIRHRLFRQ TLQIRALSPA YRRVEQSGIR 201 PRRHLRRLGR DTDPVPPPRA HRTIRRPHRLQRTAAKPARY FGQRLLDAGG 251 KIRHRTRLAD W*

Further work revealed the following gonococcal DNA sequence <SEQ ID873>:

1 ATGAAACACA TCCACATTAT CGGTATCGGC GGCACGTTTA TGGGCGGGAT 51 TGCCGCCATTGCCAAAGAAG CCGGGTTCAA AGTCAGCGGT TGCGACGCGA 101 AGATGTATCC GCCGATGAGCACCCAGCTCG AAGCCTTGGG CATAGGCGTA 151 CACGAAGGCT TCGATGCCGC GCAGTTGGAAGAATTTCAAG CCGATATTTA 201 CGTCATCGGC AATGTCGCCA GGCGCGGGAT GGATGTGGTCGAGGCGATTT 251 TGAACCGTGG GCTGCCTTAT ATTTCCGGCC CGCAATGGCT GGCTGAAAac301 GTGCtgcacc atcaTTGGgt ACTCGGCGTG GcagggaCGC ACGGcaaAac 351gaccaCcGcg tCCATGCTCG CCTGGGTCTT GGAATATGCC GGACTCGCGC 401 CGGGCTTCCTCATCGGCGGt gtaccggaAA ATTTCGGCGT TTCCGCCCGC 451 CTACCGCAAA CGCCGCGTCAAGACCCGAAC AGCAAATCGC CGTTTTTCGT 501 CATCGAAGCC GACGAATACG ACACCGCCTTTTTCGACAAA CGCTCCAAAT 551 TCGTGCATTA TCGCCCGCGT ACCGCCGTGT TGAACAATCTGGAATTCGAC 601 CACGCCGACA TCTTCGCCGA CTTGGGCGCG ATACAGACCC AGTTCCACCA651 CCTCGTGCGC ACCGTACCAT CCGAAGGCCT CATCGTCTGC AACGGACAGC 701AGCAAAGCCT GCAAGATACT TTGGACAAAG GCTGCTGGAC GCCGGTGGAA 751 AAATTCGGCACCGGACACGG CTGGCAGATT GGTGAAGTCA ATGCCGACGG 801 CTCGTTCGAC GTATTGCTTGACGGCAAAAA AGCCGGACAC GTCGCATGGG 851 ATTTGATGGG CGGACACAAC CGCATGAACGCGCTCGCCGT CATCGCTGCC 901 GCACGCCATG CCGGAGTCGA TGTTCAGACG GCCTGCGAAGCCTTGGGTGC 951 GTTTAAAAAC GTCAAACGCC GCATGGAAAT CAAAGGCACG GCAAACGGCA1001 TCACCGTTTA CGACGATTTC GCCCACCACC CGACCGCCAT CGAAACCACG 1051ATTCAAGGTT TGCGCCAACG TGTCGGCGGC GCGCGCATCC TCGCCGTCCT 1101 CGAGCCGCGTTCCAACACCA TGAAACTCGG CACGATGAAG TCCGCCCTGC 1151 CCGCAAGCCT CAAAGAAGCCGACCAAGTGT TCTGCTACGC CGGCGGCGCG 1201 GACTGGGACG TTGCCGAAGC CCTCGCGCCTTTGGGCTGCA GGCTGCGCGT 1251 CGGTAAAGAT TTCGATACCT TCGTTGCCGA AATTGTGAAAAACGCCCGAA 1301 CCGGCGACCA TATTTTGGTG ATGAGCAACG GCGGTTTCGG CGGAATACAC1351 ACCAAACTGC TGGACGCTTT GAGATAG

This corresponds to the amino acid sequence <SEQ ID 874; ORF132ng-1>:

  1 MKHIHIIGIG GTFMGGIAAI AKEAGFKVSG CDAKMYPPMS TQLEALGIGV  51HEGFDAAQLE EFQADIYVIG NVARRGMDVV EAILNRGLPY ISGPQWLAEN 101 VLHHHWVLGVAGTHGKTTTA SMLAWVLEYA GLAPGFLIGG VPENFGVSAR 151 LPQTPRQDPN SKSPFFVIEADEYDTAFFDK RSKFVHYRPR TAVLNNLEFD 201 HADIFADLGA IQTQFHHLVR TVPSEGLIVCNGQQQSLQDT LDKGCWTPVE 251 KFGTGHGWQI GEVNADGSFD VLLDGKKAGH VAWDLMGGHNRMNALAVIAA 301 ARHAGVDVQT ACEALGAFKN VKRRMEIKGT ANGITVYDDF AHHPTAIETT351 IQGLRQRVGG ARILAVLEPR SNTMKLGTMK SALPASLKEA DQVFCYAGGA 401DWDVAEALAP LGCRLRVGKD FDTFVAEIVK NARTGDHILV MSNGGFGGIH 451 TKLLDALR*

ORF132ng-1 and ORF132-1 show 93.2% identity in 458 aa overlap:

In addition, ORF132ng-1 is homologous to a hypothetical E. coli protein:

pir||S556459 hypothetical protein o457 - Escherichia coli >gi|537075 (U14003)ORF_o457 [Escherichia coli] >gi|1790660 (AE000494). hypothetical 48.5 kD proteinin fbp-pmba intergenic region [Escherichia coli] Length = 457 Score =474 bits (1207), Expect = e−133 Identities = 249/439 (56%), Positives =294/439 (66%), Gaps = 13/439 (2%)Query:  22 KEAGFKVSGCDAKMYPPMSTQLEALGIGVHEGFDAAQLEEFQADIYVIGNVARRGMDVVE  81           ++ G +V+G DA +YPPMST LE  GI + +G+DA+QLE  Q D+ +IGN   RG    VESbjct:  21 RQLGHEVTGSDANVYPPMSTLLEKQGIELIQGYDASQLEP-OPDLVIIGNAMTRGNPCVE  79Query:  82 AILNRGLPYISGPQWLAENVLHHHWVLGVAGTHGKTTTASMLAWVLEYAGLAPGFLIGGV 141           A+L  ++PY+SGPQWL  +VL   WVL VAGTHGKTTTA M  W+LE  G   PGF+IGGVSbjct:  80 AVLEKNIPYMSGPQWLHDFVLADRWVLAVAGTHGKTTTAGMATWILEQCGYKPGFVIGGV 139Query: 142 PENFGVSARLPQTPRQDPNSKSPFFVIEADEYDTAFFDKRSKFVHYRPRTAVLNNLEFDH 201           P NF VSA L          +S FFVIEADEYD AFFDKRSKFVHY PRT +LNNLEFDHSbjct: 140 PGNFEVSAHL---------GESDFFVIEADEYDCAFFDKRSKFVHYCPRTLILNNLEFDH 190Query: 202 ADIFADLGAIQTQFHHLVRTVPSEGLIVCNGQQQSLQDTLDKGCWTPVEKFGTGHGWQIG 261           ADIF DL AIQ QFHHLVR VP +G I+      +L+ T+  GCW+  E  G     WQSbjct: 191 ADIFDDLKAIQKQFHHLVRIVPGOGRIIWPENDINLKQTMAMGCWSEQELVGEQGHWQAK 250Query: 262 EVNADGS-FDVLLDGKKAGHVAWDLMGGHNRMNALAVIAAARHAGVDVQTACEALGAFKN 320           ++  D S ++VLLDG+K G V W L+G HN  N L  IAAARH GV    A   ALG+FNSbjct: 251 KLTTDASEWEVLLDGEKVGEVKWSLVGEHNMHNGLMAIAAARHVGVAPADAANALGSFIN 310Query: 321 VKRRMEIKGTANGITVYDDFAHHPTAIETTIQGLRQRVGG-ARILAVLEPRSNTMKLGTM 379            +RR+E++G ANG+TVYDDFAHHPTAI  T+  LR +VGG  ARI+AVLEPRSNTMK+GSbjct: 311 ARRRLELRGEANGVTVYDDFAHHPTAILATLAALRGKVGGTARIIAVLEPRSNTMKMGIC 370Query: 380 KSALPASLKEADQVF-CYAGGADWDVAEALAPLGCRLRVGKDFDTFVAEIVKNARTGDHI 438           K  L  SL  AD+VF        W VAE             D DT    +VK A+ GDHISbjct: 371 KDDLAPSLGRADEVFLLQPAHIPWQVAEVAEACVQPAHWSGDVDTLADMVVKTAQPGDHI 430Query: 439 LVMSNGGFGGIHTKLLDAL                                          457           LVMSNGGFGGIH KLLD LSbjct: 931 LVMSNGGFGGIHQKLLDGL                                          999

Based on this analysis, it was predicted that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

ORF132-1 (26.4 kDa) was cloned in pET and pGex vectors and expressed inE. coli, as described above. The products of protein expression andpurification were analyzed by SDS-PAGE. FIG. 20A shows the results ofaffinity purification of the His-fusion protein, and FIG. 20B shows theresults of expression of the GST-fusion in E. coli. Purified His-fusionprotein was used to immunise mice, whose sera were used for FACSanalysis (FIG. 20C) and ELISA (positive result). These experimentsconfirm that ORF132 is a surface-exposed protein, and that it is auseful immunogen.

Example 103

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 875>

   1 ..CCGGGCTATT ACGGCTCGGA TGACGAATTT AAGCGGGCAT TCGGAGAAAA  51   CTCGCCGACA TmCAAGAAAC ATTGCAACCG GAGCTGCGGG ATTTATGAAC 101   CCGTATTGAA AAAATACGGC AAAAAGCGCG CCAACAACCA TTCGGTCAGC 151   ATTAGTGCGG ACTTCGGCGA TTATTTCATG CCGTTCGCCA GCTATTCGCG 201   CACACACCGT ATGCCCAACA TCCAAGAAAT GTATTTTTCC CAAATCGGCG 251   ACTCCGGCGT TCACACCGCC TTAAAACCAG AGCGCGCAAA CACTTGGCAA 301   TTTGGCTTCr ATACCTATAA AAAAGGATTG TTAAAACAAG ATGATACATT 351   AGGATTAAAA CTGGTCGGCT ACCGCAGCCG CATCGACAAC TACATCCACA 401   ACGTTTACGG GAAATGGTGG GATTTGAACG GGGATATTCC GAGCTGGGTC 451   AGCAGCACCG GGCTTGCCTA CACCATCCAA CATCGCrATT TCAwAGACAA 501   AGTGCATCAA nnnnnnnnnn nnnnnnnnnn nnnnTACGAT TATGGGCGTT 551   TTTTCACCAA CCTTTCTTAC GCCTATCAAA AAAGCACGCA ACCGACCAAC 601   TTCAGCGATG CGAGCGAATC GCCCAACAAT GCGTCCAAAG AAGACCAACT 651   CAAACAAGGT TATGGGTTGA GCAGGGTTTC CGCCCTGCCG CGAGATTACG 701   GACGTTTGGA AGTCGGTACG CGCTGGTTGG GCAACAAACT GACTTTGGGC 751   GGCGCGATGC GCTATTTCGG CAAGAGCATC CGCGCGACGG CTGAAGAACG 801   CTATATCGAC GGCACCAACG GGGGAAATAC CAGCAATTTC CGGCAACTGG 851   GCAAGCGTTC CATCAAACAA ACCGAAACTC TTGCCCGCCA GCCTTTGATT 901   TTwGATTTTa ACGCCGCTTA CGAGCCGAAG AAAAACCTTA TTTTCCGCGC 951   CGAAGTCAAA AATCTGTTCG ACAGGCGTTA TATCGATCCG CTCGATGCGG1001   GCAATGATGC GGCAAC.GAG CGTTATTACA GCTCGTTCGA CCCGAAAGAC1051   AAGGACrrAG ACGTAACGTG TAATGCTGAT AAAACGTTGT GCaACGGCAA1101   ATACGGCGGC ACAAGCAAAA GCGTATTGAC CAATTTTGCA CGCGGACGCA1151   CCTTTTTgAT GACGATGAGC TACAAGTTTT AA

This corresponds to the amino acid sequence <SEQ ID 876; ORF133>:

  1 ..PGYYGSDDEF KRAFGENSPT XKKHCNRSCG IYEPVLKKYG KKRANNHSVS  51  ISADFGDYFM PFASYSRTHR MPNIQEMYFS QIGDSGVHTA LKPERANTWQ 101  FGFXTYKKGL LKQDDTLGLK LVGYRSRIDN YIHNVYGKWW DLNGDIPSWV 151  SSTGLAYTIQ HRXFXDKVHQ XXXXXXXXYD YGRFFTNLSY AYQKSTQPTN 201  FSDASESPNN ASKEDQLKQG YGLSRVSALP RDYGRLEVGT RWLGNKLTLG 251  GAMRYFGKSI RATAEERYID GTNGGNTSNF RQLGKRSIKQ TETLARQPLI 301  XDFNAAYEPK KNLIFRAEVK NLFDRRYIDP LDAGNDAAXE RYYSSFDPKD 351  KDXDVTCNAD KTLCNGKYGG TSKSVLTNFA RGRTFLMTMS YKF*

Further work revealed the further partial DNA sequence <SEQ ID 877>:

1 GAGGCGCAGA TACAGGTTTT GGAAGATGTG CACGTCAAGG CGAAGCGCGT 51 ACCGAAAGACAAAAAAGTGT TTACCGATGC GCGTGCCGTA TCGACCCGTC 101 AGGATATATT CAAATCCAGCGAAAACCTCG ACAACATCGT ACGCAGCATC 151 CCCGGTGCGT TTACACAGCA AGATAAAAGCTCGGGCATTG TGTCTTTGAA 201 TATTCGCGGC GACAGCGGGT TCGGGCGGGT CAATACGATGGTGGACGGCA 251 TCACGCAGAC CTTTTATTCG ACTTCTACCG ATGCGGGCAG GGCAGGCGGT301 TCATCTCAAT TCGGTGCATC TGTCGACAGC AATTTTATTG CCGGACTGGA 351TGTCGTCAAA GGCAGCTTCA GCGGCTCGGC AGGCATCAAC AGCCTTGCCG 401 GTTCGGCGAATCTGCGGACT TTAGGCGTGG ATGACGTCGT TCAGGGCAAT 451 AATACCTACG GCCTGCTGCTAAAAGGTCTG ACCGGCACCA ATTCAACCAA 501 AGGTAATGCG ATGGCGGCGA TAGGTGCGCGCAAATGGCTG GAAAGCGGAG 551 CATCTGTCGG TGTGCTTTAC GGGCACAGCA GGCGCAGCGTGGCGCAAAAT 601 TACCGCGTGG GCGGCGGCGG GCAGCACATC GGAAATTTTG GCGCGGAATA651 TTTGGAACGG CGCAAGCAGC GATATTTTGT ACAAGAGGGT GCTTTGAAAT 701TCAATTCCGA CAGCGGAAAA TGGGAGCGGG ATTTACAAAG GCAACAGTGG 751 AAATACAAGCCGTATAAAAA TTACAACAAC CAAGAACTAC AaAAATACAT 801 CGAAGAGCAT GACAAAAGCTGGCGGGAAAA CCTg.CaCCG CAATACGACA 851 TTACCCCCAT CGATCCGTCC AGCCTGAAGCAGCAGTCGGC AGGCAATCTG 901 TTTAAATTGG AATACGACGG CGTATTCAAT AAATACACGGCGCAATTTCG 951 CGATTTAAAC ACCAAAATCG GCAGCCGCAA AATCATCAAC CGCAATTATC1001 AGTTCAATTA CGGTTTGTCT TTGAACCCGT ATACCAACCT CAATCTGACC 1051GCAGCCTACA ATTCGGGCAG GCAGAAATAT CCGAAAGGGT CGAAGTTTAC 1101 AGGCTGGGGGCTTTTAAAGG ATTTTGAAAC CTACAACAAC GCGAAAATCC 1151 TCGACCTCAA CAACACCGCCACCTTCCGGC TGCCCCGCGA AACCGAGTTG 1201 CAAACCACTT TGGGCTTCAA TTATTTCCACAACGAATACG GCAAAAACCG 1251 CTTTCCTGAA GAATTGGGGC TGTTTTTCGA CGGTCCTGATCAGGACAACG 1301 GGCTTTATTC CTATTTGGGG CGGTTTAAGG GCGATAAAGG GCTGCTGCCC1351 CAAAAATCAA CCATTGTCCA ACCGGCCGGC AGCCAATATT TCAACACGTT 1401CTACTTCGAT GCCGCGCTCA AAAAAGACAT TTACCGCTTA AACTACAGCA 1451 CCAATACCGTCGGCTACCGT TTCGGCGGCG AATATACGGG CTATTACGGC 1501 TCGGATGACG AATTTAAGCGGGCATTCGGA GAAAACTCGC CGACATACAA 1551 GAAACATTGC AACCGGAGCT GCGGGATTTATGAACCCGTA TTGAAAAAAT 1601 ACGGCAAAAA GCGCGCCAAC AACCATTCGG TCAGCATTAGTGCGGACTTC 1651 GGCGATTATT TCATGCCGTT CGCCAGCTAT TCGCGCACAC ACCGTATGCC1701 CAACATCCAA GAAATGTATT TTTCCCAAAT CGGCGACTCC GGCGTTCACA 1751CCGCCTTAAA ACCAGAGCGC GCAAACACTT GGCAATTTGG CTTCAATACC 1801 TATAAAAAAGGATTGTTAAA ACAAGATGAT ACATTAGGAT TAAAACTGGT 1851 CGGCTACCGC AGCCGCATCGACAACTACAT CCACAACGTT TACGGGAAAT 1901 GGTGGGATTT GAACGGGGAT ATTCCGAGCTGGGTCAGCAG CACCGGGCTT 1951 GCCTACACCA TCCAACATCG CAATTTCAAA GACAAAGTGCACAAACACGG 2001 TTTTGAGTTG GAGCTGAATT ACGATTATGG GCGTTTTTTC ACCAACCTTT2051 CTTACGCCTA TCAAAAAAGC ACGCAACCGA CCAACTTCAG CGATGCGAGC 2101GAATCGCCCA ACAATGCGTC CAAAGAAGAC CAACTCAAAC AAGGTTATGG 2151 GTTGAGCAGGGTTTCCGCCC TGCCGCGAGA TTACGGACGT TTGGAAGTCG 2201 GTACGCGCTG GTTGGGCAACAAACTGACTT TGGGCGGCGC GATGCGCTAT 2251 TTCGGCAAGA GCATCCGCGC GACGGCTGAAGAACGCTATA TCGACGGCAC 2301 CAACGGGGGA AATACCAGCA ATTTCCGGCA ACTGGGCAAGCGTTCCATCA 2351 AACAAACCGA AACTCTTGCC CGCCAGCCTT TGATTTTTGA TTTTTACGCC2401 GCTTACGAGC CGAAGAAAAA CCTTATTTTC CGCGCCGAAG TCAAAAATCT 2451GTTCGACAGG CGTTATATCG ATCCGCTCGA TGCGGGCAAT GATGCGGCAA 2501 CGCAGCGTTATTACAGCTCG TTCGACCCGA AAGACAAGGA CGAAGACGTA 2551 ACGTGTAATG CTGATAAAACGTTGTGCAAC GGCAAATACG GCGGCACAAG 2601 CAAAAGCGTA TTGACCAATT TTGCACGCGGACGCACCTTT TTGATGACGA 2651 TGAGCTACAA GTTTTAA

This corresponds to the amino acid sequence <SEQ ID 878; ORF133-1>:

  1 EAQIQVLEDV HVKAKRVPKD KKVFTDARAV STRQDIFKSS ENLDNIVRSI  51PGAFTQQDKS SGIVSLNIRG DSGFGRVNTM VDGITQTFYS TSTDAGRAGG 101 SSQFGASVDSNFIAGLDVVK GSFSGSAGIN SLAGSANLRT LGVDDVVQGN 151 NTYGLLLKGL TGTNSTKGNAMAAIGARKWL ESGASVGVLY GHSRRSVAQN 201 YRVGGGGQHI GNFGAEYLER RKQRYFVQEGALKFNSDSGK WERDLQRQQW 251 KYKPYKNYNN QELQKYIEEH DKSWRENLXP QYDITPIDPSSLKQQSAGNL 301 FKLEYDGVFN KYTAQFRDLN TKIGSRKIIN RNYQFNYGLS LNPYTNLNLT351 AAYNSGRQKY PKGSKFTGWG LLKDFETYNN AKILDLNNTA TFRLPRETEL 401QTTLGFNYFH NEYGKNRFPE ELGLFFDGPD QDNGLYSYLG RFKGDKGLLP 451 QKSTIVQPAGSQYFNTFYFD AALKKDIYRL NYSTNTVGYR FGGEYTGYYG 501 SDDEFKRAFG ENSPTYKKHCNRSCGIYEPV LKKYGKKRAN NHSVSISADF 551 GDYFMPFASY SRTHRMPNIQ EMYFSQIGDSGVHTALKPER ANTWQFGFNT 601 YKKGLLKQDD TLGLKLVGYR SRIDNYIHNV YGKWWDLNGDIPSWVSSTGL 651 AYTIQHRNFK DKVHKHGFEL ELNYDYGRFF TNLSYAYQKS TQPTNFSDAS701 ESPNNASKED QLKQGYGLSR VSALPRDYGR LEVGTRWLGN KLTLGGAMRY 751FGKSIRATAE ERYIDGTNGG NTSNFRQLGK RSIKQTETLA RQPLIFDFYA 801 AYEPKKNLIFRAEVKNLFDR RYIDPLDAGN DAATQRYYSS FDPKDKDEDV 851 TCNADKTLCN GKYGGTSKSVLTNFARGRTF LMTMSYKF*

Computer analysis of this amino acid sequence gave the followingresults:

Homology with the Probable TonB-Dependent Receptor HI121 of H.influenzae (Accession Number U32801)

ORF133 and HI121 show 57% aa identity in 363aa overlap:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF133 shows 90.8% identity over a 392aa overlap with an ORF (ORF133a)from strain A of N. meningitidis:

A partial ORF133a nucleotide sequence <SEQ ID 879> is:

1 AAAGACAAAA AAGTGTTTAC CGATGCGCGT GCCGTATCGA CCCGTCAGGA 51 TATATTCAAATCCANCGAAA ACCTCGACAA CATCGTACGC ANCATCCCCG 101 GTGCGTTTAC ACANCAANATAAAAGCTCGG GCNTTGTGTC TTTGAATATT 151 CGCNGCGACA GCGGGTTCGG GCGGGTCAATACNATGGTNG ACGGCATCAC 201 NCANACCTTT TATTCGACTT CTACCGATGC GGGCAGGGCAGGCGGTTCAT 251 CTCAATTCGG TGCATCTGTC GACAGCAATT TTATNGCCGG ACTGGATGTC301 GTCAAAGGCA GCTTCAGCGG CTCGGCAGGC ATCAACAGCC TTGCCGGTTC 351GGCGAATCTG CGGACTTTAN GCGTGGATGA TGTCGTTCAG GGCAATANTA 401 CNTACGGCCTGCTGCTAAAA GGTCTGACCG GCACCAATTC AACCAAAGGT 451 AATGCGATGG CGGCGATAGGTGCGCGCAAA TGGCTGGAAA GCGGAGCATC 501 TGTCGGTGTG CTTTACGGGC ACAGCAGGCGCAGCGTGGCG CAAAATTACC 551 GCGTGGGCGG CGGCGGGCAG CACATCGGAA ATTTTGGCGCGGAATATCTG 601 GAACGACGCA AGCAACGATA TTTTGAGCAA GAAGGCGGGT TGAAATTCAA651 TTCCAACAGC GGAAAATGGG AGCGGGATTT CCAAAAGTCG TACTGGAAAA 701CCAAGTGGTA TCAAAAATAC GATGCCCCCC AAGAACTGCA AAAATACATC 751 GAAGGTCATGATAAAAGCTG GCGGGAAAAC CTGGCGCCGC AATACGACAT 801 CACCCCCATC GATCCGTCCAGCCTGAAGCN GCAGTCGGCA GGCAACCTGT 851 TTAAATTGGA ATACGACGGC GTATTCAATAAATACACGGC GCAATTTCGC 901 GATTTAAACA CCAAAATCGG CAGCCGCAAA ATCATCAACCGCAATTATCA 951 ATTCAATTAC GGTTTGTCTT TGAACCCGTA TACCAACCTC AATCTGACCG1001 CAGCCTACAA TTCGGGCAGG CAGAAATATC CGAAAGGGTC GAAGTTTACA 1051GGCTGGGGGC TTTTNAAAGA TTTTGAAACC TACAACAACG CAAAAATCCT 1101 CGACCTCANCAACACCTCCA CCTTCCGGCT GCCCCGTGAA ACCGAGTTGC 1151 AAACCACTTT GGGCTTCAATTATTTCCACA ACGAATACGG CAAAAACCGC 1201 TTTCCTGAAG AATTGGGGCT GTTTTTCGACGGTCCGGATC ANGACAACGG 1251 GCTTTATTCC TATTTGGGGC GGTTTAAGGG CGATAAAGGGCTGCTGCCCC 1301 AAAAATCAAC CATTGTCCAA CCGGCCGGCA GCCAATATTT CAACACGTTC1351 TACTTCGATG CCGCGCTCAA AAAAGACATT TACCGCTTAA ACTACAGCAC 1401CAATACCGTC GGCTACCGTT TCGGCGGCNA ATATACGGGC TATTACNGCT 1451 CGGATGACGAATTTAAGCGG GCATTCGGAG AAAACTCGCC GACATACANG 1501 AAACATTGCA ACCAGAGCTGCGGAATTTAT GAACCCGTAT TGAAAAAATA 1551 CGGCAAAAAG CGCGCCAACA ACCATTCGGTCAGCATTAGT GCGGACTTCG 1601 GCGATTATTT CATGCCGTTC GCCAGCTATT CGCGCACACACCGTATGCCC 1651 AACATCCAAG AAATGTATTT TTCCCAAATC GGCGACTCCG GCGTTCACAC1701 CGCCTTAAAA CCAGAGCGCG CAAACACTTG GCAATTTGGC TTCAATACCT 1751ATAAAAAAGG ATTGTTAAAA CAAGATGATA TATTAGGATT AAAACTGGTC 1801 GGCTACCGCAGCCGCATCGA CNACTACATC CACAACGTTT ACGGGAAATG 1851 GTGGGATTTG AACGGGAATATTCCGAGCTG GGTCAGCAGC ACCGGGCTTG 1901 CCTACACCAT CCAACACCGC AATTTCAAAGACAAAGTGCA CAAACACGGT 1951 TTTGAGTTGG AGCTGAATTA CGATTATNGG CGTTTTTTCACCAACCTTTC 2001 TTACGCCTAT CAAAAAAGCA CGCAACCGAC CAACTTCAGC GATGCGAGCG2051 AATCGCCCAA CAATGCGTCC AAAGAAGACC AACTCAAACA AGGTTATGGG 2101TTGAGCAGGG TTTCCGCCCT GCCGCGAGAT TACGGACGTT TGGAAGTCGG 2151 TACGCGCTGGTTGGGCAACA AACTGACTTT GGGCGGCGCG ATGCGCTATT 2201 TCGGCAAGAG CATCCGCGCGACGGCTGAAG AACGCTATAT CGACGNCACC 2251 AATGGGGNAN NTACCAGCAA TTTCCGGCAACTGGGCAAGC GTTCCATCAN 2301 ACAAACCGAA ACCCTTGCCC GCCAGCCTTT GATTTTTGATTTNTACGCCG 2351 CTTACGAGCC GAAGAAAAAN CTTATTTTCC GCGCCGAAGT CAAAAATCTG2401 TTCGACAGGC GTTATATCGA TCCGCTCGAT GCGGGCAATG ATGCGGCAAC 2451GCAGCGTTAT TACAGTTCGT TCGACCCGAA AGACAAGGAC GAAGAAGTAA 2501 CGTGTAATGATGATAACACG TTATGCAACG GCAAATACGG CGGCACAAGC 2551 AAAAGCGTAT TGACCAATTTTGCACGCGGA CNCACCTTTT TGATAACGAT 2601 GAGCTACAAG TTTTAA

This encodes a protein having (partial) amino acid sequence <SEQ ID880>:

  1 KDKKVFTDAR AVSTRQDIFK SXENLDNIVR XIPGAFTXQX KSSGXVSLNI  51RXDSGFGRVN TMVDGITXTF YSTSTDAGRA GGSSQFGASV DSNFXAGLDV 101 VKGSFSGSAGINSLAGSANL RTLXVDDVVQ GNXTYGLLLK GLTGTNSTKG 151 NAMAAIGARK WLESGASVGVLYGHSRRSVA QNYRVGGGGQ HIGNFGAEYL 201 ERRKQRYFEQ EGGLKFNSNS GKWERDFQKSYWKTKWYQKY DAPQELQKYI 251 EGHDKSWREN LAPQYDITPI DPSSLKXQSA GNLFKLEYDGVFNKYTAQFR 301 DLNTKIGSRK IINRNYQFNY GLSLNPYTNL NLTAAYNSGR QKYPKGSKFT351 GWGLXKDFET YNNAKILDLX NTSTFRLPRE TELQTTLGFN YFHNEYGKNR 401FPEELGLFFD GPDXDNGLYS YLGRFKGDKG LLPQKSTIVQ PAGSQYFNTF 451 YFDAALKKDIYRLNYSTNTV GYRFGGXYTG YYXSDDEFKR AFGENSPTYX 501 KHCNQSCGIY EPVLKKYGKKRANNHSVSIS ADFGDYFMPF ASYSRTHRMP 551 NIQEMYFSQI GDSGVHTALK PERANTWQFGFNTYKKGLLK QDDILGLKLV 601 GYRSRIDXYI HNVYGKWWDL NGNIPSWVSS TGLAYTIQHRNFKDKVHKHG 651 FELELNYDYX RFFTNLSYAY QKSTQPTNFS DASESPNNAS KEDQLKQGYG701 LSRVSALPRD YGRLEVGTRW LGNKLTLGGA MRYFGKSIRA TAEERYIDXT 751NGXXTSNFRQ LGKRSIXQTE TLARQPLIFD XYAAYEPKKX LIFRAEVKNL 801 FDRRYIDPLDAGNDAATQRY YSSFDPKDKD EEVTCNDDNT LCNGKYGGTS 851 KSVLTNFARG XTFLITMSYK F*

ORF133a and ORF133-1 show 94.3% identity in 871 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF133 shows 92.3% identity over 392 aa overlap with a predicted ORF(ORF133ng) from N. gonorrhoeae:

The complete length ORF133ng nucleotide sequence <SEQ ID 881> ispredicted to encode a protein having amino acid sequence <SEQ ID 882>:

  1 MRSSFRLKPI CFYLMGVMLY HHSYAEDAGR AGSEAQIQVL EDVHVKAKRV  51PKDKKVFTDA RAVSTRQDVF KSGENLDNIV RSIPGAFTQQ DKSSGIVSLN 101 IRGDSGFGRVNTMVDGITQT FYSTSTDAGR AGGSSQFGAS VDSNFIAGLD 151 VVKGSFSGSA GINSLAGSANLRTLGVDDVV QGNNTYGLLL KGLTGTNSTK 201 GNAMAAIGAR KWLESGASVG VLYGHSRRGVAQNYRVGGGG QHIGNFGEEY 251 LERRKQQYFV QEGGLKFNAG SGKWERDLQR QYWKTKWYKKYEDPQELQKY 301 IEEHDKSWRE NLAPQYDITP IDPSGLKQQS AGNLLNLEYD GVFNKYTAQF351 RDLNTRIGSR KIINRNYQFN YGLSLNPYTN LNLTAAYNSG RQKYPKGAKF 401TGWGLLKDFE TYNNAKILDL NNTATFRLPR ETELQTTLGF NYFHNEYGKN 451 RFPEELGLFFDGPDQDNGLY SYLGRFKGDK GLLPQKSTIV QPAGSQYFNT 501 FYFDAALKKD IYRLNYSTNAINYRFGGEYT GYYGSENEFK RAFGENSPAY 551 KEHCDPSCGL YEPVLKKYGK KRANNHSVSISADFGDYFMP FAGYSRTHRM 601 PNIQEMYFSQ IGDSGVHTAL KPERANTWQF GFNTYKKGLLKQDDILGLKL 651 VGYRSRIDNY IHNVYGKWWD LNGDIPSWVG STGLAYTIRH RNFKDKVHKH701 GFELELNYDY GRFFTNLSYA YQKSTQPTNF SDASESPNNA SKEDQLKQGY 751GLSRVSALPR DYGRLEVGTR WLGNKLTLGG AMRYFGKSIR ATAEERYIDG 801 TNGGNTSNVRQLGKRSIKQT ETLARQPLIF DFYAAYEPKK NLIFRAEVKN 851 LFDRRYIDPL DAGNDAATQRYYSSFDPKDK DEDVTCNADK TLCNGKYGGT 901 SKSVLTNFAR GRTFLMTMSY KF*

A variant was also identified, being encoded by the gonococcal DNAsequence <SEQ ID 883>:

   1 ATGAGATCTT CTTTCCGGTT GAAGCCGATT TGTTTTTATC TTATGGGTGT  51 TATGCTATAT CATCATAGTT ATGCCGAAGA TGCAGGGCGC GCGGGCAGCG 101 AGGCGCAGAT ACAGGTTTTG GAAGATGTGC ACGTCAAGGC GAAGCGCGTA 151 CCGAAAGACA AAAAAGTGTT TACCGATGCG CGTGCCGTAT CGACCCGTca 201 gGATGTGTTC AAATCCGGCG AAAACCTCGA CAACATCGTA CGCAGCATAC 251 CCGGTGCGTT TACACAGCAA GATAAAAGCT CGGGCATTGT GTCTTTGAAT 301 ATTCCCGGCG ACAGCGGGTT CGGGCGGGTC AATACGATGG TGGACGGCAT 351 CACGCAGACC TTTTATTCGA CTTCTACCGA TGCGGGCAGG GCAGGCGGTT 401 CATCTCAATT CGGTGCATCT GTCGACAGCA ATTTTATTGC CGGACTGGAT 451 GTCGTCAAAG GCAGCTTCAG CGGCTCGGCA GGCATCAACA GCCTTGCCGG 501 TTCGGCGAAT CTGCGGACTT TAGGCGTGGA TGACGTCGTT CAGGGCAATA 551 ATACCTACGG CCTGCTGCTA AAAGGTCTGA CCGGCACCAA TTCAACCAAA 601 GGTAATGCGA TGGCGGCGAT AGGTGCGCGC AAATGGCTGG AAAGCGGAGC 651 GTCTGTCGGT GTGCTTTACG GGCACAGCAG GCGCGGCGTG GCGCAAAATT 701 ACCGCGTGGG CGGCGGCGGG CAGCACATCG GAAATTTTGG TGAAGAATAT 751 CTGGAACGGC GCAAACAGCA ATATTTTGTA CAAGAGGGTG GTTTGAAATT 801 CAATGCCGGC AGCGGAAAAT GGGAACGGGA TTTGCAAAGG CAATACTGGA 851 AAACAAAGTG GTATAAAAAA TACGAAGACC CCCAAGAACT GCAAAAATAC 901 ATCGAAGAGC ATGATAAAAG CTGGCGGGAA AACCTGGCGC CGCAATACGA 951 CATCACCCCC ATCGATCCGT CCGGCCTGAA GCAGCAGTCG GCAGGCAATC1001 TGTTTAAATT GGAATACGAC GGCGTATTCA ATAAATACAC GGCGCAATTT1051 CGCGATTTAA ACACCAGAAT CGGCAGCCGC AAAATCATCA ACCGCAATTA1101 TCAATTCAAT TACGGTTTGT CTTTGAACCC GTATACCAAC CTCAATCTGA1151 CCGCAGCCTA CAATTCGGGC AGGCAGAAAT ATCCGAAAGG GGCGAAGTTT1201 ACAGGCTGGG GGCTTTTAAA AGATTTTGAA ACCTACAACA ACGCGAAAAT1251 CCTCGACCTC AACAACACCG CCACCTTCCG GCTGCCCCGC GAAACCGAGT1301 TGCAAACCAC TTTGGGCTTC AATTATTTCC ACAACGAATA CGGCAAAAAC1351 CGCTTTCCTG AAGAATTGGG GCTGTTTTTC GACGGTCCTG ATCAGGACAA1401 CGGGCTTTAT TCCTATTTGG GGCGGTTTAA GGGCGATAAA GGGCTGTTGC1451 CTCAAAAATC AACCATTGTC CAACCGGCCG GCAGCCAATA TTTCAACACG1501 TTCTACTTCG ATGCCGCGCT CAAAAAAGAC ATTTACCGCT TAAACTACAG1551 CACCAATGCA ATCAACTACC GTTTCGGCGG CGAATATACG GGCTATTACG1601 GCTCGGAAAA CGAATTTAAG CGGGCATTCG GAGAAAACTC GCCGGCATAC1651 AAGGAACATT GCGACCCGAG CTGCGGGCTT TATGAACCCG TATTGAAAAA1701 ATACGGCAAA AAGCGCGCCA ACAACCATTC GGTCAGCATT AGTGCGGACT1751 TCGGCGATTA TTTCATGCCG TTCGCCGGCT ATTCGCGCAC ACACCGTATG1801 CCCAACATCC AAGAAATGTA TTTTTCCCAA ATCGGCGACT CCGGCGTTCA1851 CACCGCCTTA AAACCAGAGC GCGCAAACAC TTGGCAATTT GGCTTCAATA1901 CCTATAAAAA AGGATTGTTA AAACAAGATG ATATATTAGG ATTGAAACTG1951 GTCGGCTACC GCAGCCGCAT TGACAACTAC ATCCACAACG TTTACGGGAA2001 ATGGTGGGAT TTGAACGGGG ATATTCCGAG CTGGGTCGGC AGCACCGGGC2051 TTGCCTACAC CATCCGACAC CGCAATTTCA AAGACAAAGT GCACAAACAC2101 GGTTTTGAGC TGGAGCTGAA TTACGATTAT GGGCGTTTTT TCACCAACCT2151 TTCTTACGCC TATCAAAAAA GCACGCAACC GACCAATTTC AGCGATGCGA2201 GCGAATCGCC CAACAATGCC tccaaAGAAG ACCAACTCAA ACAAGGTTAT2251 GGGCTGAGCA GGGTTTCCGC CCTGCCGCGA GATTACGGAC GTTTGGAAGT2301 CGGTACGCGC TGGTTGGGCA ACAAACTGAC TTTGGGCGGC GCGAtgCGCT2351 ATTTCGGCAA GAGCATCCGC GCGACGGCTG AAGAACGCTA TATCGACGGC2401 ACCAACGGGG GAAATACCAG CAATGTCCGG CAACTGGGCA AGCGTTCCAT2451 CAAACAAACC GAAACCCTTG CCCGACAGCC TTTGATTTTT GATTTTTACG2501 CCGCTTACGA GCCGAAGAAA AACCTTATTT TCCGCGCCGA AGTCAAAAAC2551 CTGTTCGACA GGCGTTATAT CGATCCGCTC GATGCGGGCA ATGATGCGGC2601 AACGCAGCGT TATTACAGCT CGTTCGACCC GAAAGACAAG GACGAAGACG2651 TAACGTGTAA TGCTGATAAA ACGTTGTGCA ACGGCAAATA CGGCGGCACA2701 AGCAAAAGCG TATTGACCAA TTTCGCACGC GGACGCACCT TCTTGATGAC2751 GATGAGCTAC AAGTTTTAA

This corresponds to the amino acid sequence <SEQ ID 884; ORF133ng-1>:

  1 MRSSFRLKPI CFYLMGVMLY HHSYAEDAGR AGSEAQIQVL EDVHVKAKRV 51 PKDKKVFTDA RAVSTRQDVF KSGENLDNIV RSIPGAFTQQ DKSSGIVSLN101 IRGDSGFGRV NTMVDGITQT FYSTSTDAGR AGGSSQFGAS VDSNFIAGLD151 VVKGSFSGSA GINSLAGSAN LRTLGVDDVV QGNNTYGLLL KGLTGTNSTK201 GNAMAAIGAR KWLESGASVG VLYGHSRRGV AQNYRVGGGG QHIGNFGEEY251 LERRKQQYFV QEGGLKFNAG SGKWERDLQR QYWKTKWYKK YEDPQELQKY301 IEEHDKSWRE NLAPQYDITP IDPSGLKQQS AGNLFKLEYD GVFNKYTAQF351 RDLNTRIGSR KIINRNYQFN YGLSLNPYTN LNLTAAYNSG RQKYPKGAKF401 TGWGLLKDFE TYNNAKILDL NNTATFRLPR ETELQTTLGF NYFHNEYGKN451 RFPEELGLFF DGPDQDNGLY SYLGRFKGDK GLLPQKSTIV QPAGSQYFNT501 FYFDAALKKD IYRLNYSTNA INYRFGGEYT GYYGSENEFK RAFGENSPAY551 KEHCDPSCGL YEPVLKKYGK KRANNHSVSI SADFGDYFMP FAGYSRTHRM601 PNIQEMYFSQ IGDSGVHTAL KPERANTWQF GFNTYKKGLL KQDDILGLKL651 VGYRSRIDNY IHNVYGKWWD LNGDIPSWVG STGLAYTIRH RNFKDKVHKH701 GFELELNYDY GRFFTNLSYA YQKSTQPTNF SDASESPNNA SKEDQLKQGY751 GLSRVSALPR DYGRLEVGTR WLGNKLTLGG AMRYFGKSIR ATAEERYIDG801 TNGGNTSNVR QLGKRSIKQT ETLARQPLIF DFYAAYEPKK NLIFRAEVKN851 LFDRRYIDPL DAGNDAATQR YYSSFDPKDK DEDVTCNADK TLCNGKYGGT901 SKSVLTNFAR GRTFLMTMSY KF*

ORF133ng-1 and ORF133-1 show 96.2% identity in 889 aa overlap:

In addition, ORF133ng-1 is homologous to a TonB-dependent receptor in H.influenzae:

sp|P45114|YC17_HAEIN PROBABLE TONB-DEPENDENT RECEPTOR HI1217 PRECURSOR >gi|1075372|pir||G64110 transferrin binding protein 1 precursor (tbp1)homolog - Haemophilus influenzae (strain Rd KW20) >gi|1574147 (U32801)transferrin binding protein 1 precursor (tbp1) [Haemophilus influenzae]Length = 913 Score = 930 bits (2377), Expect = 0.0 Identities =476/921 (51%), Positives = 619/921 (66%), Gaps = 72/921 (7%) Query: 38QVLEDVHVKAKRVPKDKKVFTDARAVSTRQDVFKSGENLDNIVRSIPGAFTQQDKSSGIV 97 + L  +V  K +  DKK FT+A+A STR++VFK  + +D ++RSIPGAFTQQDK SG+V Sbjct: 29ETLGQIDVVEKVISNDKKPFTEAKAKSTRENVFKETQTIDQVIRSIPGAFTQQDKGSGVV 88 Query:98 SLNIRGDSGFGRVNTMVDGITQTFYSTSTDAGRAGGSSQFGASVDSNFIAGLDVVKGSFS 157S+NIRG++G GRVNTMVDG+TQTFYST+ D+G++GGSSQFGA++D NFIAG+DV K +FS Sbjct: 89SVNIRGENGLGRVNTMVDGVTQTFYSTALDSGQSGGSSQFGAAIDPNFIAGVDVNKSNFS 148 Query:158 GSAGINSLAGSANLRTLGVDDVVQXXXXXXXXXXXXXXXXXXXXXAMAAIGARKWLESGA 217G++GIN+LAGSAN RTLGV+DV+                       M     RKWL++G Sbjct: 149GASGINALAGSANFRTLGVNDVITDDKPFGIILKGMTGSNATKSNFMTMAAGRKWLDNGG 208 Query:218 SVGVLYGHSRRGVAQNYRVGGGGQHIGNFGEEYLERRKQQYFVQEGGLKFNAGSGKWERD 277 VGV+YG+S+R V+Q+YR+ GGG+ + + G++ L + K+ YF +  G   N   G+W  D Sbjct: 209YVGVVYGYSQREVSQDYRI-GGGERLASLGQDILAKEKEAYF-RNAGYILNP-EGQWTPD 265 Query:278 LQRQYWK-----------TKWY--------------------KKYEDPQELQK---YIEE 303L +++W              +Y                    KK +D ++LQK    IEE Sbjct: 266LSKKHWSCNKPDYQKNGDCSYYRIGSAAKTRREILQELLTNGKKPKDIEKLQKGNDGIEE 325 Query:304 HDKSWRENLAPQYDITPIDPSGLKQQSAGNLFKLEYDGVFNKYTAQFRDLNTRIGSRKII 363 DKS+  N   QY + PI+P  L+ +S  +L K EY        AQ R L+ +IGSRKI Sbjct: 326TDKSFERN-KDQYSVAPIEPGSLQSRSRSHLLKFEYGDDHQNLGAQLRTLDNKIGSRKIE 384 Query:364 NRNYQFNYGLSLNPYTNLNLTAAYNSGRQKYPKGAKFTGWGLLKDFETYNNAKILDLNNT 423NRNYQ NY  + N Y +LNL AA+N G+  YPKG  F GW +     T N A I+D+NN+ Sbjct: 385NRNYQVNYNFNNNSYLDLNLMAAHNIGKTIYPKGGFFAGWQVADKLITKNVANIVDINNS 444 Query:424 ATFRLPRETELQTTLGFNYFHNEYGKNRFPEELGLFFDGPDQDNGLYSY--LGRFKGDKG 481 TF LP+E +L+TTLGFNYF NEY KNRFPEEL LF++    D GLYS+   GR+ G K Sbjct: 445HTFLLPKEIDLKTTLGFNYFTNEYSKNRFPEELSLFYNDASHDQGLYSHSKRGRYSGTKS 504 Query:482 LLPQKSTIVQPAGSQYFNTFYFDAALKKDIYRLNYSTNAINYRFGGEYTGYYGSENEFKR 541LLPQ+S I+QP+G Q F T YFD AL K IY LNYS N  +Y F GEY GY Sbjct: 505LLPQRSVILQPSGKQKFKTVYFDTALSKGIYHLNYSVNFTHYAFNGEYVGY--------- 555 Query:542 AFGENSPAYKEHCDPSCGLYEPVLKKYGKKRANNHSVSISADFGDYFMPFAGYSRTHRMP 601   EN+   +        + EP+L K G K+A NHS ++SA+  DYFMPF  YSRTHRMP Sbjct: 556---ENTAGQQ--------INEPILHKSGHKKAFNHSATLSAELSDYFMPFFTYSRTHRMP 604 Query:602 NIQEMYFSQIGDSGVHTALKPERANTWQFGFNTYKKGLLKQDDILGLKLVGYRSRIDNYI 661NIQEM+FSQ+ ++GV+TALKPE+++T+Q GFNTYKKGL  QDD+LG+KLVGYRS I NYI Sbjct: 605NIQEMFFSQVSNAGVNTALKPEQSDTYQLGFNTYKKGLFTQDDVLGVKLVGYRSFIKNYI 664 Query:662 HNVYGKWWDLNGDIPSWVGSTGLAYTIRHRNFKDKVHKHGFELELNYDYGRFFTNLSYAY 721HNVYG WW     +P+W  S G  YTI H+N+K  V K G ELE+NYD GRFF N+SYAY Sbjct: 665HNVYGVWW--RDGMPTWAESNGFKYTIAHQNYKPIVKKSGVELEINYDMGRFFANVSYAY 722 Query:722 QKSTQPTNFSDASESPNNASKEDQLKQGYGLSRVSALPRDYGRLEVGTRWLGNKLTLGGA 781 Q++QPTN++DAS  PNNAS+ED LKQGYGLSRVS LP+DYGRLE+GTRW   KLTLG A Sbjct: 723QRTNQPTNYADASPRPNNASQEDILKQGYGLSRVSMLPKDYGRLELGTRWFDQKLTLGLA 782 Query:782 MRYFGKSIRATAEERYIDGTNGGNTSNVRQLGKRSIKQTETLARQPLIFDFYAAYEPKKN 841RY+GKS RAT EE YI+G+     + +R+    ++K+TE + +QP+I D + +YEP K+ Sbjct: 783ARYYGKSKRATIEEEYINGSR-FKKNTLRRENYYAVKKTEDIKKQPIILDLHVSYEPIKD 841 Query:842 LIFRAEVKNLFDRRYIDPLDAGNDAATQRYYSSFDPKDKDEDVTCNADKTLCNGKYGGTS 901LI +AEV+NL D+RY+DPLDAGNDAA+QRYYSS      +  + C  D + C    GG+ Sbjct: 842LIIKAEVQNLLDKRYVDPLDAGNDAASQRYYSSL-----NNSIECAQDSSAC----GGSD 892 Query:902 KSVLTNFARGRTFLMTMSYKF 922 K+VL NFARGRT++++++YKF Sbjct: 893KTVLYNFARGRTYILSLNYKF 913

The underlined motif in the gonococcal protein (also present in themeningococcal protein) is predicted to be an ATP/GTP-binding site motifA (P-loop), and the analysis suggests that these proteins from N.meningitidis and N. gonorrhoeae, and their epitopes, could be usefulantigens for vaccines or diagnostics, or for raising antibodies.

Example 104

The following partial DNA sequence was identified in N. meningitidis<SEQ ID 885>

1 ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG TTATGGCGGT 51TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT GAAATCCTGT 101ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG GGAAATGCTG 151GGCTACACCG CCCTCAAAAT GCCCGCCCGC GCCTACGAAC TGATTCCCCT 201CGCCGTCCTT ATCGGCGGAC TGGTCTCCCT CAGCCAGCTT GCCGCCGGCA 251GCGAACTGAC CGTCATCAAA GCCAGCGGCA TGAGCACCAA AAAGCTGCTG 301TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA CCGTCGCGCT 351CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA AACATCAAAG 401CCGCCGCCAT CAACGGCAAA ATCAGCACCG GCAATACCGG CCTTTGGCTG 451AAAGAAAAAA ACAGCGTGAT CAATGTGCGC GAAATGTTGC CCGACCAT..

This corresponds to the amino acid sequence <SEQ ID 886; ORF112>:

1 MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG KGSYGIWEML 51GYTALKMPAR AYELIPLAVL IGGLVSLSQL AAGSELTVIK ASGMSTKKLL 101LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK ISTGNTGLWL 151KEKNSVINVR EMLPDH...

Further work revealed further partial nucleotide sequence <SEQ ID 887>:

1 ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG TTATGGCGGT 51TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT GAAATCCTGT 101ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG GGAAATGCTG 151gGCTACACCG CCCTCAAAAT GCCCGCCCGC GCCTACGAAC TGATTCCCCT 201CGCCGTCCTT ATCGGCGGAC TGGTCTCCCT CAGCCAGCTT GCCGCCGGCA 251GCGAACTGAC CGTCATCAAA GCCAGCGGCA TGAGCACCAA AAAGCTGCTG 301TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA CCGTCGCGCT 351CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA AACATCAAAG 401CCGCCGCCAT CAACGGCAAA ATCAGCACCG GCAATACCGG CCTTTGGCTG 451AAAGAAAAAA ACAGCrTkAT CAATGTGCGC GAAATGTTGC CCGACCATAC 501GCTTTTGGGC ATCAAAATTT GGGCGCGCAA CGATAAAAAC GAATTGGCAG 551AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGG CAGTTGGCAG 601TTGAAAAACA TCCGCCGCAG CACGCTTGGC GAAGACAAAG TCGAGGTCTC 651TATTGCGGCT GAAGAAAACT GGCCGATTTC CGTCAAACGC AACCTGATGG 701ACGTATTGCT CGTCAAACCC GACCAAATGT CCGTCGGCGA ACTGACCACC 751TACATCCGCC ACCTCCAAAA CAACAGCCAA AACACCCGAA TCTACGCCAT 801CGCATGGTGG CGCAAATTGG TTTACCCCGC CGCAGCCTGG GTGATGGCGC 851TCGTCGCCTT TGCCTTTACC CCGCAAACCA CCCGCCACGG CAATATGGGC 901TTAAAACTCT TCGGCGGCAT CTGTsTCGGA TTGCTGTTCC ACCTTGCCGG 951ACGGCTCTTT GGGTTTACCA GCCAACTCGG...

This corresponds to the amino acid sequence <SEQ ID 888; ORF112-1>:

1 MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG KGSYGIWEML 51GYTALKMPAR AYELIPLAVL IGGLVSLSQL AAGSELTVIK ASGMSTKKLL 101LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK ISTGNTGLWL 151KEKNSXINVR EMLPDHTLLG IKIWARNDKN ELAEAVEADS AVLNSDGSWQ 201LKNIRRSTLG EDKVEVSIAA EENWPISVKR NLMDVLLVKP DQMSVGELTT 251YIRHLQNNSQ NTRIYAIAWW RKLVYPAAAW VMALVAFAFT PQTTRHGNMG 301LKLFGGICXG LLFHLAGRLF GFTSQL...

Computer analysis of this amino acid sequence predicts two transmembranedomains and gave the following results:

Homology with a Predicted ORF from N. meningitidis (Strain A)

ORF112 shows 96.4% identity over a 166aa overlap with an ORF (ORF112a)from strain A of N. meningitidis:

The ORF112a nucleotide sequence <SEQ ID 889> is:

1 ATGAACCTGA TTTCACGTTA CATCATCCGT CAAATGGCGG TTATGGCGGT 51TTACGCGCTC CTTGCCTTCC TCGCTTTGTA CAGCTTTTTT GAAATCCTGT 101ACGAAACCGG CAACCTCGGC AAAGGCAGTT ACGGCATATG GGAAATGNTG 151GGNTACACCG CCCTCAAAAT GNCCGCCCGC GCCTACGAAC TGATGCCCCT 201CGCCGTCCTT ATCGGCGGAC TGGTCTCTNT CAGCCAGCTT GCCGCCGGCA 251GCGAACTGAN CGTCATCAAA GCCAGCGGCA TGAGCACCAA AAAGCTGCTG 301TTGATTCTGT CGCAGTTCGG TTTTATTTTT GCTATTGCCA CCGTCGCGCT 351CGGCGAATGG GTTGCGCCCA CACTGAGCCA AAAAGCCGAA AACATCAAAG 401CCGCGGCCAT CAACGGCAAA ATCAGTACCG GCAATACCGG CCTTTGGCTG 451AAAGAAAAAA ACAGCATTAT CAATGTGCGC GAAATGTTGC CCGACCATAC 501CCTGCTGGGC ATTAAAATCT GGGCCCGCAA CGATAAAAAC GAACTGGCAG 551AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGG CAGTTGGCAG 601TTGAAAAACA TCCGCCGCAG CACGCTTGGC GAAGACAAAG TCGAGGTCTC 651TATTGCGGCT GAAGAAAANT GGCCGATTTC CGTCAAACGC AACCTGATGG 701ACGTATTGCT CGTCAAACCC GACCAAATGT CCGTCGGCGA ACTGACCACC 751TACATCCGCC ACCTCCAAAN NNACAGCCAA AACACCCGAA TCTACGCCAT 801CGCATGGTGG CGCAAATTGG TTTACCCCGC CGCAGCCTGG GTGATGGCGC 851TCGTCGCCTT TGCCTTTACC CCGCAAACCA CCCGCCACGG CAATATGGGC 901TTAAAANTCT TCGGCGGCAT CTGTCTCGGA TTGCTGTTCC ACCTTGCCGG 951NCGGCTCTTC NGGTTTACCA GCCAACTCTA CGGCATCCCG CCCTTCCTCG 1001NCGGCGCACT ACCTACCATA GCCTTCGCCT TGCTCGCCGT TTGGCTGATA 1051CGCAAACAGG AAAAACGCTA A

This encodes a protein having the amino acid sequence <SEQ ID 890>:

1 MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG KGSYGIWEMX 51GYTALKMXAR AYELMPLAVL IGGLVSXSQL AAGSELXVIK ASGMSTKKLL 101LILSQFGFIF AIATVALGEW VAPTLSQKAE NIKAAAINGK ISTGNTGLWL 151KEKNSIINVR EMLPDHTLLG IKIWARNDKN ELAEAVEADS AVLNSDGSWQ 201LKNIRRSTLG EDKVEVSIAA EEXWPISVKR NLMDVLLVKP DQMSVGELTT 251YIRHLQXXSQ NTRIYAIAWW RKLVYPAAAW VMALVAFAFT PQTTRHGNMG 301LKXFGGICLG LLFHLAGRLF XFTSQLYGIP PFLXGALPTI AFALLAVWLI 351 RKQEKR*

ORF112a and ORF112-1 show 96.3% identity in 326 aa overlap:

Homology with a Predicted ORF from N. gonorrhoeae

ORF112 shows 95.8% identity over 166aa overlap with a predicted ORF(ORF112ng) from N. gonorrhoeae:

The complete length ORF112ng nucleotide sequence <SEQ ID 891> is:

1 ATGAACCTGA TTTCACGTTA CATCATCCGC CAAATGGCGG TTATGGCGGT 51 TTACGCGCTCCTTGCCTTCC TCGCTTTGTA CAGCTTTTTT GAAATCCTGT 101 ACGAAACCGG CAACCTCGGCAAAGGCAGTT ACGGCATATG GGAAATGCTG 151 GGCTACACCG CCCTCAAAAT GCCCGCCCGCGCCTACGAAC TCATGCCCCT 201 CGCCGTCCTC ATCGGCGGAC TGGCCTCTCT CAGCCAGCTTGCCGCCGGCA 251 GCGAACTGGC CGTCATCAAA GCCAGCGGCA TGAGCACCAA AAAGCTGCTG301 TTGATTCTGT CTCAGTTCGG TTTTATTTTT GCTATTGCCG CCGTCGCGCT 351CGGCGAATGG GTTGCGCCCA CGCTGAGCCA AAAAGCCGAA AACATCAAag 401 cCGCCGCCAttaacggCAAA ATCAGCAccg gcAATACCGG CCTTTggcTG 451 AAAGAAAAAa ccAGCATTATCAATGTGcGc GGAATGTTGC CCGACCATAC 501 GCTTTTGGGC ATCAAAATTT GGGCGCGCAACGATAAAAAC GAATTGGCAG 551 AGGCAGTGGA AGCCGATTCC GCCGTTTTGA ACAGCGACGGCAGCTGGCAG 601 TTGAAAAACA TCCGCCGCAG CATCATGGGT ACAGACAAAA TCGAAACATC651 cgCCGCCGCC GAAGAAACTT gGCCGATTGC CGTCAGACGC AACCTGATGG 701ACGTATTGCT CGTCAAGCCC GACCAAATGT CCGTCGGCGA GCTGACCACC 751 TACATCCGCCACCTCCAAAA CAACAGCCAA AACACCCAAA TCTACGCCAT 801 CGCATGGTGG CGTAAACTCGTTTACCCCGT CGCCGCATGG GTCATGGCGC 851 TCGTTGCCTT CGCCTTTACG CCGCAAACCACGCGCCACGG CAATATGGGC 901 TTAAAACTCT TCGGCGGCAT CTGTCTCGGA TTGCTGTTCCACCTTGCCGG 951 CAGGCTCTTC GGGTTTACCA GCCAACTCTA CGGCACCCCA CCCTTCCTCG1001 CCGGCGCACT GCCTACCATA GCCTTCGCCT TGCTCGCTGT TTGGCTGATA 1051CGCAAACAGG AAAAACGTTG A

This encodes a protein having amino acid sequence <SEQ ID 892>:

1 MNLISRYIIR QMAVMAVYAL LAFLALYSFF EILYETGNLG KGSYGIWEML 51GYTALKMPAR AYELMPLAVL IGGLASLSQL AAGSELAVIK ASGMSTKKLL 101LILSQFGFIF AIAAVALGEW VAPTLSQKAE NIKAAAINGK ISTGNTGLWL 151 KEKTSIINVRGMLPDHTLLG IKIWARNDKN ELAEAVEADS AVLNSDGSWQ 201 LKNIRRSIMG TDKIETSAAAEETWPIAVRR NLMDVLLVKP DQMSVGELTT 251 YIRHLQNNSQ NTQIYAIAWWRKLVYPVAAW VMALVAFAFT PQTTRHGNMG 301 LKLFGGICLG LLFHLAGRLF GFTSQLYGTPPFLAGALPTI AFALLAVWLI

-   -   351 RKQEKR*

ORF112ng and ORF112-1 show 94.2% identity in 326 aa overlap:

This analysis suggests that these proteins from N. meningitidis and N.gonorrhoeae, and their epitopes, could be useful antigens for vaccinesor diagnostics, or for raising antibodies.

It will be appreciated that the invention has been described by means ofexample only, and that modifications may be made whilst remaining withinthe spirit and scope of the invention.

TABLE I PCR primers ORF Primer Sequence Restriction sites ORF 1 ForwardCGCGGATCCGCTAGC-GGACACACTTATTTCGG BamHI-NheI ReverseCCCGCTCGAG-CCAGCGGTAGCCTAATT XhoI ORF 2 ForwardGCGGATCCCATATG-TTTGATTTCGGTTTGGG BamHI-NdeI ReverseCCCGCTCGAG-GACGGCATAACGGCG XhoI ORF 2-1 ForwardGCGGATCCCATATG-TTTGATTTCGGTTTGGG BamHI-NdeI ReverseCCCGCTCGAG-TGATTTACGGACGCGCA XhoI ORF 4 ForwardGCGGATCCCATATG-TGCGGAGGTCAAAAAGAC BamHI-NdeI ReverseCCCGCTCGAG-TTTGGCTGCGCCTTC XhoI ORF 5 ForwardGGAATTCCATATGGCCATGG-TGGAAGGCGCACAACC NdeI-NcoI ForwardCGGGATCC-ATGGAAGGCGCACAAC BamHI Reverse CCCGCTCGAG-GACTGTGCAAAAACGG XhoIORF 6 Forward CGCGGATCCCATATG-ACCCGTCAATCTCTGCA BamHI-NdeI ReverseCCCGCTCGAG-TGCGCCGAACACTTTC XhoI ORF 7 ForwardCGCGGATCCGCTAGC-GCGCTGCTTTTTGTTCC BamHI-NheI ReverseCCCGCTCGAG-TTTCAAAATATATTTGCGGA XhoI ORF 8 ForwardGCGGATCCCATATG-GCTCAACTGCTTCGTAC BamHI-NdeI ReverseCCCGCTCGAG-AGCAGGCTTTGGCGC XhoI ORF 9 ForwardCGCGGATCCCATATG-CCGAAGGAAGTCGGAAA BamHI-NdeI ReverseCCCGCTCGAG-TTTCCGAGGTTTTCGGG XhoI ORF 10 ForwardGCGGATCCCATATG-GACACAAAAGAAATCCTC BamHI-NdeI ReverseCCCGCTCGAG-TAATGGGAAACCTTGTTTT XhoI ORF 11 ForwardGCGGATCCCATATG-GCGGTCAACCTCTACG BamHI-NdeI ReverseCCCGCTCGAG-GGAAACGACTTCGCC XhoI ORF 13 ForwardCGCGGATCCCATATG-GCTCTGCTTTCCGCGC BamHI-NdeI ReverseCCCGCTCGAG-AGGGTGTGTGATAATAAG XhoI ORF 15 ForwardGGAATTCCATATGGCCATGG-GCGGGACACTGACAG NdeI-NcoI ForwardCGGGATCC-TGCGGGACACTGACAGG BamHI Reverse CCCGCTCGAG-AGGTTGGCCTTGTCTATGXhoI ORF 17 Forward GGAATTCCATATGGCCATGG-TTGCCGGCCTGTTCG NdeI-NcoIForward CGGGATCC-ATTGCCGGCCTGTTCG BamHI ReverseCCCGCTCGAG-AAGCAGGTTGTACAGC XhoI ORF 18 ForwardGCGGATCCCATATG-ATTTTGCTGCATTTGGAT BamHI-NdeI ReverseCCCGCTCGAG-TCTTCCAATTTCTGAAAGC XhoI ORF 19 ForwardGGAATTCCATATGGCCATGG -TCGCCAGTGTTTTTACC NdeI-NcoI ForwardCGGGATCC-TTCGCCAGTGTTTTTACCG BamHI Reverse CCCGCTCGAG-GGTGTTTTTGAAGCTGCCXhoI ORF 20 Forward GGAATTCCATATGGCCATGG -TCGGCGCGGGTATG NdeI-NcoIForward CGGGATCC-TTCGGCGCGGGTATG BamHI ReverseCCCGCTCGAG-CGGCGAGCGAGAGCA XhoI ORF 22 ForwardGGAATTCCATATGGCCATGG-TGATTAAAATCAAAAAAGGTCT NdeI-NcoI ForwardCGGGATCC-ATGATTAAAATCAAAAAAGGTCTAAACC BamHI ReverseCCCGCTCGAG-ATTATGATAGCGGCCC XhoI ORF 23 ForwardCGCGGATCCCATATG-GATGTTTCTGTTTCAGAC BamHI-NdeI ReverseCCCGCTCGAG-TTTAAACCGATAGGTAAACG XhoI ORF 24 ForwardGGAATTCCATATGGCCATGG- TGATGCCGGAAATGGTG NdeI-NcoI ForwardCGGGATCC-ATGATGCCGGAAATGGTG BamHI Reverse CCCGCTCGAG-TGTCAGCGTGGCGCAXhoI ORF 25 Forward GCGGATCCCATATG-TATCGCAAACTGATTGC BamHI-NdeI ReverseCCCGCTCGAG-ATCGATGGAATAGCCG XhoI ORF 26 ForwardGCGGATCCCATATG -CAGCTGATCGACTATTC BamHI-NdeI ReverseCCCGCTCGAG-GACATCGGCGCGTTTT XhoI ORF 27 ForwardGGAATTCCATATGGCCATGG-AGACCTATTCTGTTTA NdeI-NcoI Forward CGGGATCC-CAGACCTATTCTGTTTATTTTAATC BamHI Reverse CCCGCTCGAG-GGGTTCGATTAAATAACCATXhoI ORF 28 Forward GGAATTCCATATGGCCATGG-ACGGCTGTACGTTGATGT NdeI-NcoIForward CGGGATCC-AACGGCTGTACGTTGATG BamHI ReverseCCCGCTCGAG-TTTGTCAGAGGAATTCGCG XhoI ORF 29 ForwardGCGGATCCCATATG -AACGGTTTGGATGCCCG BamHI-NdeI ForwardCGCGGATCCGCTAGC-AACGGTTTGGATGCCCG BamHI-NheI ReverseCCCGCTCGAG-TTTGTCTAAGTTCCTGATATG XhoI ORF 32 ForwardCGCGGATCCCATATG-AATACTCCTCCTTTTG BamHI-NdeI ReverseCCCGCTCGAG-GCGTATTTTTTGATGCTTTG XhoI ORF 33 ForwardGCGGATCCCATATG -ATTGATAGGGATCGTATG BamHI-NdeI ReverseCCCGCTCGAG-TTGATCTTTCAAACGGCC XhoI ORF 35 ForwardGCGGATCCCATATG-TTCAGAGCTCAGCTT BamHI-NdeI ForwardCGCGGATCCGCTAGC-TTCAGAGCTCAGCTT BamHI-NheI ReverseCCCGCTCGAG-AAACAGCCATTTGAGCGA XhoI ORF 37 ForwardGCGGATCCCATATG-GATGACGTATCGGATTTT BamHI-NdeI ReverseCCCGCTCGAG-ATAGCCCGCTTTCAGG XhoI ORF 58 ForwardCGCGGATCCGCTAGC-TCCGAACGCGAGTGGAT BamHI-NheI ReverseCCCGCTCGAG-AGCATTGTCCAAGGGGAC XhoI ORF 65 ForwardGGAATTCCATATGGCCATGG -TGCTGTATCTGAATCAAG NdeI-NcoI ForwardCGGGATCC-TTGCTGTATCTGAATCAAGG BamHI Reverse CCCGCTCGAG-CCGCATCGGCAGACAXhoI ORF 66 Forward GCGGATCCCATATG-TACGCATTTACCGCCG BamHI-NdeI ReverseCCCGCTCGAG-TGGATTTTGCAGAGATGG XhoI ORF 72 Forward CGCGGATCCCATATG-AATGCAGTAAAAATATCTGA BamHI-NdeI Reverse CCCGCTCGAG-GCCTGAGACCTTTGCAAXhoI ORF 73 Forward GCGGATCCCATATG-AGATTTTTCGGTATCGG BamHI-NdeI ReverseCCCGCTCGAG-TTCATCTTTTTCATGTTCG XhoI ORF 75 Forward GCGGATCCCATATG-TCTGTCTTTCAAACGGC BamHI-NdeI Reverse CCCGCTCGAG-TTTGTTTTTGCAAGACAG XhoIORF 76 Forward GATCAGCTAGCCATATG-AAACAGAAAAAAACCGC NheI-NdeI ReverseCGGGATCC-TTACGGTTTGACACCGTT BamHI ORF 79 ForwardCGCGGATCCCATATG-GTTTCCGCCGCCG BamHI-NdeI ReverseCCCGCTCGAG-GTGCTGATGCGCTTCG XhoI ORF 83 ForwardGCGGATCCCATATG-AAAACCCTGCTGCTGC BamHI-NdeI ReverseCCCGCTCGAG-GCCGCCTTTGCGGC XhoI ORF 84 ForwardGCGGATCCCATATG-GCAGAGATCTGTTTG BamHI-NdeI ReverseCCCGCTCGAG-GTTTGCCGATCCGACCA XhoI ORF 85 Forward CGCGGATCCCATATG-GCGGTTTGGGGCGGA BamHI-NdeI Reverse CCCGCTCGAG-TCGGCGCGGCGGGC XhoI ORF 89Forward GGAATTCCATATGGCCATGG-CCATACCTTCTTATCA NdeI-NcoI ForwardCGGGATCC-GCCATACCTTCTTATCAGAG BamHI ReverseCCCGCTCGAG-TTTTTTGCGATTAGAAAAAGC XhoI ORF 97 ForwardGCGGATCCCATATG-CATCCTGCCAGCGAAC BamHI-NdeI ReverseCCCGCTCGAG-TTCGCCTACGGTTTTTTG XhoI ORF 98 ForwardGCGGATCCCATATG-ACGGTAACTGCGG BamHI-NdeI ReverseCCCGCTCGAG-TTGTTGTTCGGGCAAATC XhoI ORF 100 ForwardGCGGATCCCATATG-TCGGGCATTTACACCG BamHI-NdeI ReverseCCCGCTCGAG-ACGGGTTTCGGCGGAA XhoI ORF 101 ForwardGCGGATCCCATATG-ATTTATCAAAGAAACCTC BamHI-NdeI ReverseCCCGCTCGAG-TTTTCCGCCTTTCAATGT XhoI ORF 102 ForwardGCGGATCCCATATG-GCAGGGCTGTTTTACC BamHI-NdeI ReverseCCCGCTCGAG-AAACGGTTTGAACACGAC XhoI ORF 103 ForwardGCGGATCCCATATG-AACCACGACATCAC BamHI-NdeI ReverseCCCGCTCGAG-CAGCCACAGGACGGC XhoI ORF 104 ForwardGCGGATCCCATATG-ACGTGGGGAACGC BamHI-NdeI ReverseCCCGCTCGAG-GCGGCGTTTGAACGGC XhoI ORF 105 ForwardGCGGATCCCATATG-ACCAAATTTCAAACCCCTC BamHI-NdeI ReverseCCCGCTCGAG-TAAACGAATGCCGTCCAG XhoI ORF 106 ForwardGCGGATCCCATATG-AGGATAACCGACGGCG BamHI-NdeI ReverseCCCGCTCGAG-TTTGTTCCCGATGATGTT XhoI ORF 109 ForwardGCGGATCCCATATG-GAAGATTTATATATAATACTCG BamHI-NdeI ReverseCCCGCTCGAG-ATCAGCTTCGAACCGAAG XhoI ORF 110 ForwardAAAGAATTC-ATGAGTAAATCCCGTAGATCTCCC EcoRI ReverseAAACTGCAG-GGAAAACCACATCCGCACTCTGCC PstI ORF 111 ForwardAAAGAATTC-GCACCGCAAAAGGCAAAAACCGCA EcoRI ReverseAAACTGCAG-TCTGCGCGTTTTCGGGCAGGGTGG PstI ORF 113 ForwardAAAGAATTC-ATGAACAAAACCCTCTATCGTGTGATTTTCAACCG EcoRI ReverseAAACTGCAG-TTACGAATGCCTGCTTGCTCGACCGTACTG PstI ORF 115 ForwardAAAGAATTC-TTGCTTGTGCAAACAGAAAAAGACGG EcoRI ReverseAAAAAAGTCGAC-CTATTTTTTAGGGGCTTTTGCTTGTTTGAAAAGCCTGCC SalI ORF 119Forward AAAGAATTC-TACAACATGTATCAGGAAAACCAATACCG EcoRI ReverseAAACTGCAG-TTATGAAAACAGGCGCAGGGCGGTTTTGCC PstI ORF 120 ForwardAAAGAATTC-GCAAGGCTACCCCAATCCGCCGTG EcoRI ReverseAAACTGCAG-CGGTTTGGCTGCCTGGCCGTTGAT PstI ORF 121 ForwardAAAGAATTC-GCCTTGGTCTGGCTGGTTTTCGC EcoRI ReverseAAACTGCAG-TCATCCGCCACCCCACCTCGGCCATCCATC PstI ORF 122 ForwardAAAAAAGTCGAC-ATGTCTTACCGCGCAAGCAGTTCTCC SalI ReverseAAACTGCAG-TCAGGAACACAAACGATGACGAATATCCGTATC PstI ORF 125 ForwardAAAGAATTC-GCGCTGTTTTTTGCGGCGGCGTAT EcoRI ReverseAAACTGCAG-CGCCGTTTCAAGACGAAAAAGTCG PstI ORF 126 ForwardAAAGAATTC-GCGGAAACGGTCGAAG EcoRI ReverseAAACTGCAG-TTAATCTTGTCTTCCGATATAC PstI ORF 127 ForwardAAAGAATTC-ATGACTGATAATCGGGGGTTTACG EcoRI ReverseAAAAAAGTCGAC-CTTAAGTAACTTGCAGTCCTTATC SalI ORF 128 ForwardAAAGAATTC-ATGCAAGCTGTCCGCTACAGGCC EcoRI ReverseAAACTGCAG-CTATTGCAATGCGCCGCCGCGGGAATGTTTGAGCAGGCG PstI ORF 129 ForwardAAAGAATTC-ATGGATTTTCGTTTTGACATTATTTACGAATACCG EcoRI ReverseAAACTGCAG-TTATTTTTTGATGAAATTTTGGGGCGG PstI ORF 130 ForwardAAAGAATTC-GCAGTACTTGCCATTCTCGGTGCG EcoRI ReverseAAACTGCAG-CTCCGGATCGTCTGTAAACGCATT PstI ORF 131 ForwardGCGGATCCCATATG-GAAATTCGGGCAATAAAAT BamHI-NdeI ReverseCCCGCTCGAG-CCAGCGGACGCGTTC XhoI ORF 132 ForwardGCGGATCCCATATG-AAAGAAGCGGGGTTTG BamHI-NdeI ReverseCCCGCTCGAG-CCAATCTGCCAGCCGT XhoI ORF 133 ForwardCGCGGATCCCATATG-GAAGATGCAGGGCGCG BamHI-NdeI ReverseCCCGCTCGAG-AAACTTGTAGCTCATCGT XhoI ORF 134 ForwardGCGGATCCCATATG-TCTGTGCAAGCAGTATTG BamHI-NdeI ReverseCCCGCTCGAG-ATCCTGTGCCAATGCG XhoI ORF 135 ForwardGCGGATCCCATATG-CCGTCTGAAAAAGCTTT BamHI-NdeI ReverseCCCGCTCGAG-AAATACCGCTGAGGATG XhoI ORF 136 ForwardCGCGGATCCGCTAGC-ATGAAGCGGCGTATAGCC BamHI-NheI ReverseCCCGCTCGAG-TTCCGAATATTTGGAACTTTT XhoI ORF 137 ForwardCGCGGATCCCATATG-GGCACGGCGGGAAATA BamHI-NdeI ReverseCCCGCTCGAG-ATAACGGTATGCCGCC XhoI ORF 138 ForwardGCGGATCCCATATG-TTTCGTTTACAATTCAGGC BamHI-NdeI ReverseCCCGCTCGAG-CGGCGTTTTATAGCGG XhoI ORF 139 ForwardGCGGATCCCATATG-GCTTTTTTGGCGGTAATG BamHI-NdeI ReverseCCCGCTCGAG-TAACGTTTCCGTGCGTTT XhoI ORF 140 ForwardGCGGATCCCATATG-TTGCCCACAGGCAGC BamHI-NdeI ReverseCCCGCTCGAG-GACGATGGCAAACAGC XhoI ORF 141 ForwardGCGGATCCCATATG-CCGTCTGAAGCAGTCT BamHI-NdeI ReverseCCCGCTCGAG-ATCTGTTGTTTTTAAAATATT XhoI ORF 142 ForwardGCGGATCCCATATG-GATAATTCTGGTAGTGAAG BamHI-NdeI ReverseCCCGCTCGAG-AAACGTATAGCCTACCT XhoI ORF 143 ForwardGCGGATCCCATATG-GATACCGCTTTGAACCT BamHI-NdeI ReverseCCCGCTCGAG-AATGGCTTCCGCAATATG XhoI ORF 144 ForwardGCGGATCCCATATG-ACCTTTTTACAACGTTTGC BamHI-NdeI ReverseCCCGCTCGAG-AGATTGTTGTTGTTTTTTCG XhoI ORF 147 ForwardGCGGATCCCATATG-TCTGTCTTTCAAACGGC BamHI-NdeI ReverseCCCGCTCGAG-TTTGTTTTTGCAAGACAG XhoI NB: restriction sites are underlinedfor ORFs 110-130, where the ORF itself carries an EcoRI site (eg. ORF122), a SalI site was used in the forward primer instead. Similarly,where the ORF carries a PstI site (eg. ORFs 115 and 127), a SalI sitewas used in the reverse primer.

TABLE II Summary of cloning, expression and purification PCR/ His-fusionGST-fusion ORF cloning expression expression Purification orf 1  + + +His-fusion orf 2  + + + GST-fusion orf 2.1  + n.d. + GST-fusion orf4  + + + His-fusion orf 5  + n.d. + GST-fusion orf 6  + + + GST-fusionorf 7  + + + GST-fusion orf 8  + n.d. n.d. orf 9  + + + GST-fusion orf10  + n.d. n.d. orf 11  + n.d. n.d. orf 13  + n.d. + GST-fusion orf15  + + + GST-fusion orf 17  + n.d. n.d. orf 18  + n.d. n.d. orf 19  +n.d. n.d. orf 20  + n.d. n.d. orf 22  + + + GST-fusion orf 23  + + +His-fusion orf 24  + n.d. n.d. orf 25  + + + His-fusion orf 26  + n.d.n.d. orf 27  + + + GST-fusion orf 28  + + + GST-fusion orf 29  + n.d.n.d. orf 32  + + + His-fusion orf 33  + n.d. n.d. orf 35  + n.d. n.d.orf 37  + + + GST-fusion orf 58  + n.d. n.d. orf 65  + n.d. n.d. orf66  + n.d. n.d. orf 72  + + n.d. His-fusion orf 73  + n.d. + n.d. orf75  + n.d. n.d. orf 76  + + n.d. His-fusion orf 79  + + n.d. His-fusionorf 83  + n.d. + n.d. orf 84  + n.d. n.d. orf 85  + n.d. + GST-fusionorf 89  + n.d. + GST-fusion orf 97  + + + GST-fusion orf 98  + n.d. n.d.orf 100 + n.d. n.d. orf 101 + n.d. n.d. orf 102 + n.d. n.d. orf 103 +n.d. n.d. orf 104 + n.d. n.d. orf 105 + n.d. n.d. orf 106 + + +His-fusion orf 109 + n.d. n.d. orf 110 + n.d. n.d. orf 111 + + n.d.His-fusion orf 113 + + n.d. His-fusion orf 115 n.d. n.d. n.d. orf119 + + n.d. His-fusion orf 120 + + n.d. His-fusion orf 121 + n.d. n.d.orf 122 + + n.d. His-fusion orf 125 + + n.d. His-fusion orf 126 + + n.d.His-fusion orf 127 + + n.d. His-fusion orf 128 + n.d. n.d. orf 129 + +n.d. His-fusion orf 130 + n.d. n.d. orf 131 + + + n.d. orf 132 + + +His-fusion orf 133 + n.d. + GST-fusion orf 134 + n.d. n.d. orf 135 +n.d. n.d. orf 136 + n.d. n.d. orf 137 + n.d. + GST-fusion orf 138 +n.d. + GST-fusion orf 139 + n.d. n.d. orf 140 + n.d. n.d. orf 141 + n.d.n.d. orf 142 + n.d. n.d. orf 143 + n.d. n.d. orf 144 + n.d. + n.d. orf147 + n.d. n.d.

1: An isolated protein comprising: (a) the amino acid sequence of SEQ IDNO: 654; or (b) an amino acid sequence having 80% or greater sequenceidentity to the amino acid sequence of SEQ ID NO: 654; or (b) a fragmentof SEQ ID NO: 654 of at least 10 contiguous amino acids in length. 2:The isolated protein of claim 1 comprising (b). 3: The isolated proteinof claim 2, wherein the amino acid sequence has 90% or greater sequenceidentity to the amino acid sequence of SEQ ID NO:
 654. 4: The isolatedprotein of claim 2, wherein the amino acid sequence has 95% or greatersequence identity to the amino acid sequence of SEQ ID NO:
 654. 5: Theisolated protein of claim 1 comprising (c). 6: A composition comprisingthe protein of any one of claims 1-5 and an adjuvant. 7: The compositionof claim 6 further comprising a pharmaceutically acceptable carrier.